The importance of physiological ecology in conservation biology.

Many of the threats to the persistence of populations of sensitive species have physiological or pathological mechanisms, and those mechanisms are best understood through the inherently integrative discipline of physiological ecology. The desert tortoise was listed under the Endangered Species Act largely due to a newly recognized upper respiratory disease thought to cause mortality in individuals and severe declines in populations. Numerous hypotheses about the threats to the persistence of desert tortoise populations involve acquisition of nutrients, and its connection to stress and disease. The nutritional wisdom hypothesis posits that animals should forage not for particular food items, but instead, for particular nutrients such as calcium and phosphorus used in building bones. The optimal foraging hypothesis suggests that, in circumstances of resource abundance, tortoises should forage as dietary specialists as a means of maximizing intake of resources. The optimal digestion hypothesis suggests that tortoises should process ingesta in ways that regulate assimilation rate. Finally, the cost-of-switching hypothesis suggests that herbivores, like the desert tortoise, should avoid switching food types to avoid negatively affecting the microbe community responsible for fermenting plants into energy and nutrients. Combining hypotheses into a resource acquisition theory leads to novel predictions that are generally supported by data presented here. Testing hypotheses, and synthesizing test results into a theory, provides a robust scientific alternative to the popular use of untested hypotheses and unanalyzed data to assert the needs of species. The scientific approach should focus on hypotheses concerning anthropogenic modifications of the environment that impact physiological processes ultimately important to population phenomena. We show how measurements of such impacts as nutrient starvation, can cause physiological stress, and that the endocrine mechanisms involved with stress can result in disease. Finally, our new syntheses evince a new hypothesis. Free molecules of the stress hormone corticosterone can inhibit immunity, and the abundance of "free corticosterone" in the blood (thought to be the active form of the hormone) is regulated when the corticosterone molecules combine with binding globulins. The sex hormone, testosterone, combines with the same binding globulin. High levels of testosterone, naturally occurring in the breeding season, may be further enhanced in populations at high densities, and the resulting excess testosterone may compete with binding globulins, thereby releasing corticosterone and reducing immunity to disease. This sequence could result in physiological and pathological phenomena leading to population cycles with a period that would be essentially impossible to observe in desert tortoise. Such cycles could obscure population fluctuations of anthropogenic origin.

Malaria surveillance--United States, 1998.

PROBLEM/CONDITION: Human malaria is caused by one or more of four species of intraerythrocytic protozoa of the genus Plasmodium (i.e., P. falciparum, P. vivax, P. ovale, or P. malariae). The protozoa are transmitted by the bite of an infective female Anopheles species mosquito. The majority of malaria infections in the United States occur among persons who have traveled to areas with endemic transmission. Cases occasionally occur that are acquired through exposure to infected blood products, by congenital transmission, or by local mosquitoborne transmission. Malaria surveillance is conducted to identify episodes of local transmission and to guide prevention recommendations for travelers. REPORTING PERIOD: Cases with an onset of symptoms during 1998. DESCRIPTION OF SYSTEM: Malaria cases confirmed by blood smear are reported to local and state health departments by health-care providers and laboratory staff members. Case investigations are conducted by local and state health departments, and reports are sent to CDC through the National Malaria Surveillance System (NMSS). This report uses NMSS data. RESULTS: CDC received reports of 1,227 cases of malaria with onsets of symptoms in 1998, among persons in the United States and its territories. This number represents a decrease of 20.5% from the 1,544 cases reported during 1997. P. falciparum, P. vivax, P. malariae, and P. ovale were identified in 42.8%, 37.8%, 3.5%, and 2.1% of cases, respectively. More than one species was present in seven patients (0.6% of total). The infecting species was not determined in 162 (13.2%) cases. Compared with reported cases in 1997, reported malaria cases acquired in Africa increased by 1.3% (n = 706); those acquired in Asia decreased by 52.1% (n = 239); and those acquired in the Americas decreased by 6.5% (n = 229). Of 636 U.S. civilians who acquired malaria abroad, 126 (19.8%) reportedly had followed a chemoprophylactic drug regimen recommended by CDC for the area to which they had traveled. Five persons became infected in the United States. One case was congenitally acquired; one was acquired by blood transfusion; and three were isolated cases that could not be epidemiologically linked to another case. Four deaths were attributed to malaria. INTERPRETATION: The 20.5% decrease in malaria cases during 1998 compared with 1997 resulted primarily from decreases in P. vivax cases acquired in Asia among non-U.S. civilians. This decrease could have resulted from local changes in disease transmission, decreased immigration from the region, decreased travel to the region, incomplete reporting from state and local health departments, or increased use of effective antimalarial chemoprophylaxis. In a majority of reported cases, U.S. civilians who acquired infection abroad had not taken an appropriate chemoprophylaxis regimen for the country where they acquired malaria. PUBLIC HEALTH ACTIONS TAKEN: Additional information was obtained from state and local health departments and clinics concerning the four fatal cases and the five infections acquired in the United States. Persons traveling to a malarious area should take a recommended chemoprophylaxis regimen and use personal protection measures to prevent mosquito bites. Any person who has been to a malarious area and subsequently develops fever or influenza-like symptoms should seek medical care immediately; the investigation should include a blood smear for malaria. Malaria infections can be fatal if not diagnosed and treated promptly. Current recommendations concerning prevention and treatment of malaria can be obtained from CDC.

Solid-phase synthesis of novel inhibitors of farnesyl transferase.

A novel diphosphate mimic, the 2,3,6-trifluoro-5-hydroxy-4-nitrophenoxy group (1), has been employed as the template in the solid-phase synthesis of novel farnesyl transferase inhibitors using the Mitsunobu reaction. The most potent inhibitor (farnesyloxy-5-hydroxy-2,3,6-trifluoro-4-nitrobenzene) displayed an IC50 of 6.3 microM versus farnesyl transferase.

Malaria surveillance--United States, 1995.

PROBLEM/CONDITION: Malaria is caused by four species of Plasmodium (i.e., P. falciparum, P. vivax, P. ovale, or P. malariae), which are transmitted by the bite of an infective female Anopheles sp. mosquito. Most malaria infections in the United States occur among persons who have traveled to areas with ongoing transmission. Occasionally, cases occur in the United States through exposure to infected blood products, by congenital transmission, or by local mosquito-borne transmission. Malaria surveillance is conducted to identify episodes of local transmission and to guide prevention recommendations for travelers. REPORTING PERIOD: Cases with onset of illness during 1995. DESCRIPTION OF SYSTEM: Malaria cases confirmed by blood smears are reported to local and/or state health departments by health-care providers and/or laboratory staff. Case investigations are conducted by local and/or state health departments, and reports are transmitted to CDC through the National Malaria Surveillance System (NMSS). Data from NMSS serve as the basis for this report. RESULTS: CDC received reports of 1,167 cases of malaria with onset of symptoms during 1995 among persons in the United States or one of its territories. This number represents an increase of 15% from the 1,014 cases reported for 1994. P. vivax, P. falciparum, P. malariae, and P. ovale were identified in 48.2%, 38.6%, 3.9%, and 2.2% of cases, respectively. More than one species was present in three patients (0.3% of total). The infecting species was not determined in 80 (6.9%) cases. The number of reported malaria cases acquired in Africa (n=519) remained approximately the same as in 1994 (n=517); cases acquired in Asia increased by 32.4% (n=335); and cases acquired in the Americas increased by 37.4 % (n=246). Of 591 U.S. civilians who acquired malaria abroad, 15.6% had followed a chemoprophylactic drug regimen recommended by CDC for the area where they had traveled. Nine patients became infected in the United States. Of these nine cases, five were congenitally acquired; one was acquired by organ transplantation; and one was acquired by a blood transfusion. For two of the nine cases, the source of infection was unknown. Six deaths were attributed to malaria. INTERPRETATION: The 15% increase in malaria cases in 1995 compared with 1994 resulted primarily from increases in cases acquired in Asia and the Americas, most notably a 100% increase in the number of cases reported from South America. This change could have resulted from local changes in disease transmission, travel patterns, reporting errors, or a decreased use of effective antimalarial chemoprophylaxis. In most reported cases, U.S. civilians who acquired infection abroad were not on an appropriate chemoprophylaxis regimen for the country where they acquired malaria. ACTIONS TAKEN: Additional information was obtained concerning the six fatal cases and the nine infections acquired in the United States. Malaria prevention guidelines were updated and distributed to health-care providers. Persons traveling to a malarious area should take the recommended chemoprophylaxis regimen and use personal protection measures to prevent mosquito bites. Any person who has been to a malarious area and who subsequently develops a fever or influenza-like symptoms should seek medical care; investigation should include a blood smear for malaria. Malaria infections can be fatal if not diagnosed and treated promptly. Recommendations concerning prevention and treatment of malaria can be obtained from CDC.

Inhibition of protein prenylation by metabolites of limonene.

The monoterpenes limonene and perillyl alcohol are undergoing clinical evaluation in cancer patients. In this paper, we report the chemical synthesis, characterisation, and quantitation in patients' plasma of a novel human metabolite of limonene, which is identified as an isomer of perillic acid. The synthesis of R-perillic acid is also described, because previous reports on the activity of perillic acid against isoprenylation enzymes refer to the S-enantiomer, although it is the R-enantiomer which is the metabolite of R-limonene. The above monoterpenes, with several related compounds, were assayed for inhibitory activity towards the isoprenylation enzymes in rat brain cytosol. Although R- and S-limonene are only weak inhibitors of the isoprenylation enzymes, their major metabolites, perillic acid and perillyl alcohol, are more potent inhibitors, with IC50 values in the low mM range. The metabolites possess greater activity towards the geranylgeranyltransferase type I enzyme than farnesyltransferase, while the novel metabolite displays IC50 values similar to those of perillic acid suggesting that it may contribute to the in vivo activity of limonene.

Monitoring for mefloquine-resistant Plasmodium falciparum in Africa: implications for travelers' health.

The effectiveness of mefloquine to prevent malaria caused by Plasmodium falciparum is influenced by the sensitivity of the malaria parasites to this drug. Concern has been raised that resistance to mefloquine may develop in sub-Saharan Africa as has been observed in Southeast Asia. Case reports, along with blood smears to confirm the diagnosis and blood samples to determine the mefloquine concentration, were provided on any Peace Corps volunteer serving in sub-Saharan Africa who was diagnosed with malaria. We defined prophylaxis failures probably due to mefloquine resistance as patients with P. falciparum malaria confirmed at the Centers for Disease Control and Prevention, reported compliance with prophylaxis, no ingestion of mefloquine between date of illness onset and date of blood drawing, and a mefloquine level > or = 620 ng/ml in blood drawn within five days of onset of illness. Between January 1, 1991 and September 6, 1996, 44 (31%) of 140 volunteers with confirmed P. falciparum had blood drawn within five days of onset of illness. Twenty-nine (66%) had not fully complied with prophylaxis. Five of 15 prophylaxis failures in four countries had mefloquine levels > or = 620 ng/ml. Failure of mefloquine prophylaxis is primarily due to noncompliance. Evidence of probable resistance to mefloquine among strains of P. falciparum was found in five Peace Corps volunteers in sub-Saharan Africa. Clusters of well-documented prophylaxis failures need to be followed-up by therapeutic in vivo studies to document parasite resistance to mefloquine. Reduced sensitivity to mefloquine does not (yet) appear to be a significant problem in sub-Saharan Africa.

Malaria surveillance -- United States, 1994.

PROBLEM/CONDITION: Malaria is caused by infection with one of four species of Plasmodium (i.e., P. falciparum, P. vivax, P. ovale, and P. malariae ), which are transmitted by the bite of an infective female Anopheles sp. mosquito. Most malarial infections in the United States occur in persons who have traveled to areas (i.e., other countries) in which disease transmission is ongoing. However, cases are transmitted occasionally through exposure to infected blood products, by congenital transmission, or by local mosquitoborne transmission. Malaria surveillance is conducted to identify episodes of local transmission and to adapt prevention recommendations. REPORTING PERIOD COVERED: Cases with onset of symptoms during 1994. DESCRIPTION OF SYSTEM: Malaria cases confirmed by blood smear are reported to local and/or state health departments by health-care providers and/or laboratories. Case investigations are conducted by local and/or state health departments, and the reports are transmitted to CDC through the National Malaria Surveillance System (NMSS), which was the source of data for this report. Numbers of cases reported through NMSS may differ from those reported through other passive surveillance systems because of differences in the collection and transmission of data. RESULTS: CDC received reports of 1,014 cases of malaria with onset of symptoms during 1994 among persons in the United States or one of its territories. This number represented a 20% decrease from the 1,275 cases reported for 1993. P. vivax, P. falciparum, P. malariae, and P. ovale accounted for 44%, 44%, 4%, and 3% of cases, respectively. More than one species was present in five persons (<1% of the total number of patients). The infecting species was not determined in 50 (5%) cases. The number of reported malaria cases in U.S. military personnel decreased by 86% (i.e., from 278 cases in 1993 to 38 cases in 1994). Of the U.S. civilians who acquired malaria during travel to foreign countries, 18% had followed a chemoprophylactic drug regimen recommended by CDC for the area to which they had traveled. Five persons became infected while in the United States; the infection was transmitted to two of these persons through transfusion of infected blood products. The remaining three cases, which occurred in Houston, Texas, were probably locally acquired mosquitoborne infections. Four deaths were attributed to malaria. INTERPRETATION: The 20% decrease in the number of malaria cases from 1993 to 1994 resulted primarily from an 86% decrease in cases among U.S. military personnel after withdrawal from Somalia. Because most malaria cases acquired in Somalia during 1993 resulted from infection with P. vivax, there was a proportionately greater decrease during 1994 in the number of cases caused by P. vivax relative to those caused by P. falciparum. ACTIONS TAKEN: Additional information was obtained concerning the four fatal cases and the five cases acquired in the United States. Malaria prevention guidelines were updated and distributed to health-care providers. Persons traveling to a geographic area in which malaria is endemic should take the recommended chemoprophylactic regimen and should use protective measures to prevent mosquito bites. Persons who have a fever or influenza-like illness after returning from a malarious area should seek medical care; medical evaluation should include a blood smear examination for malaria. Malarial infections can be fatal if not promptly diagnosed and treated. Recommendations concerning prevention and treatment of malaria can be obtained from CDC.

Malaria surveillance--United States, 1993.

PROBLEM/CONDITION: Malaria is caused by infection with one of four species of Plasmodium (P. falciparum, P. vivax, P. ovale, and P. malariae), which are transmitted by the bite of an infective female Anopheles sp. mosquito. Most malaria cases in the United States occur among persons who have traveled to areas (i.e., other countries) in which disease transmission is ongoing. However, cases are transmitted occasionally through exposure to infected blood products, by congenital transmission, or by local mosquito-borne transmission. Malaria surveillance is conducted to identify episodes of local transmission and to guide prevention recommendations. REPORTING PERIOD COVERED: Cases with onset of illness during 1993. DESCRIPTION OF SYSTEM: Malaria cases confirmed by blood smear are reported to local and/or state health departments by health-care providers and/or laboratories. Case investigations are conducted by local and/or state health departments, and the reports are transmitted to CDC. RESULTS: CDC received reports of 1,275 cases of malaria in persons in the United States and its territories who had onset of symptoms during 1993; this number represented a 40% increase over the 910 malaria cases reported for 1992. P. vivax, P. falciparum, P. ovale, and P. malariae were identified in 52%, 36%, 4%, and 3% of cases, respectively. The species was not determined in the remaining 5% of cases. The 278 malaria cases in U.S. military personnel represented the largest number of such cases since 1972; 234 of these cases were diagnosed in persons returning from deployment in Somalia during Operation Restore Hope. In New York City, the number of reported cases increased from one in 1992 to 130 in 1993. The number of malaria cases acquired in Africa by U.S. civilians increased by 45% from 1992; of these, 34% had been acquired in Nigeria. The 45% increase primarily reflected cases reported by New York City. Of U.S. civilians who acquired malaria during travel, 75% had not used a chemoprophylactic regimen recommended by CDC for the area in which they had traveled. Eleven cases of malaria had been acquired in the United States: of these cases, five were congenital; three were induced; and three were cryptic, including two cases that were probably locally acquired mosquito-borne infections. Eight deaths were associated with malarial infection. INTERPRETATION: The increase in the reported number of malaria cases was attributed to a) the number of infections acquired during military deployment in Somalia and b) complete reporting for the first time of cases from New York City. ACTIONS TAKEN: Investigations were conducted to collect detailed information concerning the eight fatal cases and the 11 cases acquired in the United States. Malaria prevention guidelines were updated and disseminated to health-care providers. Persons who have a fever or influenza-like illness after returning from a malarious area should seek medical care, regardless of whether they took antimalarial chemoprophylaxis during their stay. The medical evaluation should include a blood smear examination for malaria. Malaria can be fatal if not diagnosed and treated rapidly. Recommendations concerning prevention and treatment of malaria can be obtained from CDC.

Malaria surveillance--United States, 1992.

PROBLEM/CONDITION: Malaria is caused by one of four species of Plasmodium (i.e., P. falciparum, P. vivax, P. ovale, or P. malariae) and is transmitted by the bite of an infective female Anopheles sp. mosquito. Most malaria cases in the United States occur among persons who have traveled to areas that have ongoing transmission. However, cases are transmitted occasionally through exposure to infected blood products, by congenital transmission, or by local mosquito-borne transmission. Malaria surveillance is conducted to identify episodes of local transmission and to guide prevention recommendations. REPORTING PERIOD COVERED: Cases with onset of illness during 1992. DESCRIPTION OF SYSTEM: Malaria cases were identified at the local level (i.e., by healthcare providers or through laboratory-based surveillance). All suspected cases were confirmed by slide diagnosis and then reported to the respective state health department and to CDC. RESULTS: CDC received reports of 910 cases of malaria that had onset of symptoms during 1992 among persons in the United States and its territories. In comparison, 1,046 cases were reported for 1991, representing a decrease of 13% in 1992. P. vivax, P. falciparum, P. malariae, and P. ovale were identified in 51%, 33%, 4%, and 3% of cases, respectively. The species was not identified in the remaining 9% of cases. The number of reported malaria cases that had been acquired in Africa by U.S. civilians decreased 38%, primarily because the number of P. falciparum cases declined. Of U.S. civilians whose illnesses were diagnosed as malaria, 81% had not taken a chemoprophylactic regimen recommended by CDC. Seven patients had acquired their infections in the United States. Seven deaths were attributed to malaria. INTERPRETATION: The decrease in the number of P. falciparum cases in U.S. civilians could have resulted from a change in travel patterns, reporting errors, or increased use of more effective chemoprophylaxis regimens. ACTIONS TAKEN: Additional information was obtained concerning the seven fatal cases and the seven cases acquired in the United States. Malaria prevention guidelines were updated and disseminated to health-care providers. Persons traveling to a malaria-endemic area should take the recommended chemoprophylaxis regimen and use personal protection measures to prevent mosquito bites. Any person who has been to a malarious area and who subsequently develops a fever or influenza-like symptoms should seek medical care, which should include a blood smear for malaria. The disease can be fatal if not diagnosed and treated at an early stage of infection. Recommendations concerning prevention and treatment of malaria can be obtained from CDC.

CRP-DNA complexes: inducing the A-like form in the binding sites with an extended central spacer.

The consensus DNA sequence for binding of the Escherichia coli cyclic AMP receptor protein (CRP) has two symmetrically related inverted recognition elements TGTGA:TCACA, separated by a variable spacer, normally 6 bp long. We have shown that the CRP-cAMP complex, when bound to synthetic binding sites with an extended 8 bp spacer segment, induces an increase in the DNA circular dichroism (CD). The CD change at lambda > 275 nm agrees with the shift of approximately one helical turn of DNA into A-like form. The B-conformation is preserved for CRP binding sites similar to that in the lac and uxaCA promoters with 6 bp spacers. Another effect accompanying DNA binding is a dramatic increase of the negative CD magnitude in the spectral region of the ligand cAMP, at lambda < 272 nm. This effect is observed when CRP binds to specific sites with 6 or 8 bp spacers as well as to non-specific DNA. We reason that the A-like form arises by compressing and unwinding the DNA in CRP-DNA complexes having 8 bp central spacers. This serves to maintain a fixed length and twisting angle and is controlled by the protein's relatively rigid frame. This model is consistent with the observation that some binding sites with 6 bp spacers may also show the CD increase inherent to the sites with the extended 8 bp spacers. These 6 bp spacers are characterized by an increased twisting angle that requires their unwinding to bind to CRP. We propose that a mutual adaptation between CRP and binding sites by local untwisting and a B-->A-like transition in the DNA is of general importance and may occur in other protein-DNA complexes, such as the complex of RNA polymerase with promoter DNA.

CRP-binding sites: evidence for two structural classes with 6-bp and 8-bp spacers.

While classifying protein binding DNA sequences of the type GTGNxCAC, based on the size of Nx [Shumilov, Mol. Biologya (Engl. Transl.) 21 (1987) 168-187], we had previously found that the cyclic AMP receptor protein (CRP)-binding sites found in the Escherichia coli genome are of at least two classes: (i) those with a conventional 6-bp spacer (N6) and (ii) those with a potential 8-bp spacer (N8) [Barber and Zhurkin, J. Biomol. Struct. Dyn. 8 (1990) 213-232]. In this paper, we present the first experimental evidence that CRP binds to DNA with an N8 spacer with relatively high affinity, as measured by gel electrophoresis of CRP-DNA complexes. We have tested two types of N8 spacers: A+T-rich and G+C-rich. Compared with the affinity of CRP for a reference site with an N6 spacer, the binding strength of CRP toward an A+T-rich N8 sequence is lower and that toward a G+C-rich N8 site is comparable. Just like DNA sites with N6 spacers, those with N8 spacers utilize both halves of the symmetrical protein recognition sequences, TGTGA and TCACA. Because of the increased number of nucleotides in the N8 spacer, the two recognition sequences in DNA will have an increased distance and a helical twist between them. These would cause displacement of the two recognition sequences with respect to the two symmetrically located alpha-helices of the CRP dimer, if there is no change in the DNA conformation.(ABSTRACT TRUNCATED AT 250 WORDS)

pNiXa, a Ni(2+)-binding protein in Xenopus oocytes and embryos, shows identity to Ep45, an estrogen-regulated hepatic serpin.

A Ni(2+)-binding protein (pNiXa, 45 kD, pI 8.5) discovered in Xenopus embryos, was isolated from oocytes. Based on amino acid sequences, pNiXa belongs to the serpin superfamily and shows identity to the cDNA sequence of Ep45, an estrogen-regulated hepatic serpin that contains an (HX)n-motif found in eukaryotic transcription factors. Nondenatured pNiXa, purified by Ni-affinity chromatography, inhibited bovine alpha-chymotrypsin. The presence of pNiXa in embryos when they are susceptible to Ni2+, the high avidity of pNiXa for Ni2+, and the (HX)n-motif point to pNiXa as a molecular target of Ni(2+)-teratogenesis.

A generalized approach to detection of organophosphate resistance in mosquitoes.

Insecticide bioassays and biochemical microtitre assays were compared for detection of resistance to the organophosphate insecticides malathion and fenitrothion, using inbred laboratory strains of malaria vectors Anopheles albimanus Wiedemann, An.arabiensis Patton and An.stephensi Liston. With susceptible mosquitoes, the LT100 values determined from bioassays corresponded closely with times taken to abolish the activity of acetylcholinesterase activity in biochemical assays: approximately 2 h for malathion and 3 h for fenitrothion. Resistant strains of all three anophelines showed longer survival correlated with prolonged acetylcholinesterase activity. An.albimanus strains with insensitive acetylcholinesterase survived bioassays with discriminating doses of 1 h exposure to 5% malathion or 1% fenitrothion and were judged as resistant. It is concluded that enzyme-specific microassays provide a reliable means of detecting resistant individuals, with practical advantages over bioassays which do not reveal the resistance mechanism and require large numbers of healthy mosquitoes.

CAP binding sites reveal pyrimidine-purine pattern characteristic of DNA bending.

To investigate the intrinsic bending of DNA at sites where proteins bind, we analyzed catabolite gene activator protein (CAP) binding sites and various operators from the viewpoint of DNA bending flexibility. Theoretical conformational analysis. DNase I digestion and x-ray crystallography data indicate that bending of B-DNA is highly anisotropic and sequence-dependent. Certain dimers prefer to bend into the major groove ("major-philic") and others prefer to bend into the minor groove ("minor-philic" dimers). From these data we considered TA, CG, CA:TG and GG:CC as major-philic dimers and AT,AA:TT and GT:AC as minor-philic ones. Analysis of 31 CAP binding sites has identified strong major-philic tendencies 5-7 base pairs (bp) away from the center. In addition, we found minor-philic poly-A tracts extending 4-5 bp away from the proposed major-philic bends. Finally, to analyze the central regions we followed the lead of Shumilov and classified the DNA sites by their spacer lengths [V.Y. Shumilov, Mol. Biol. (Mosk) 21, 168-187 (1987)]. In this way, we identified two subsets of CAP binding sites: one with 6 bp between the TGTGA:TCACA consensus boxes (N6-set) and one with 8 central bp (N8-set). We discovered that the dimer at the center of an N6-set site was usually major-philic, whereas at the center of an N8-set site more often minor-philic. Analysis of phages 434, P22 lambda and trp operators revealed similar results. In conclusion, our data show that CAP binding sites have major-philic and minor-philic dimers at specific positions; the location of these dimers may facilitate wrapping of DNA around CAP. A similar pattern is seen in nucleosomes.

HIV-1 reverse transcriptase: structure predictions for the polymerase domain.

Reverse transcriptase (RT) plays an essential role in the life cycle of the human immunodeficiency viruses (HIV). A better understanding of this enzyme, and its two catalytic functions, the DNA polymerase and the RNase H, could lead to the development of new drugs that would specifically block HIV replication. The available genetic, sequence, biochemical, and immunological data on the reverse transcriptase of HIV-1 constrain the possible structure of the DNA polymerase domain. The purpose of this review is to correlate the data and to discuss, in light of that data, a model for the structure of the polymerase domain. In this model, the polymerase domain is approximately 50 to 60 A in diameter with a 20 A opening to accommodate the nucleic acid duplex. The most evolutionarily conserved region of RT (amino acids 20-190 of HIV-1 RT) is proposed to form the inner surface of the 20 A opening to which the nucleic acid hemiduplex is bound.

SequenceEditingAligner: a multiple sequence editor and aligner.

Here we present the SequenceEditingAligner system for editing multiple, aligned genetic sequences. This is an interactive multi-window color system that displays more than 3500 nucleotides or amino acids. The system handles nucleic acid or protein sequences with or without secondary structure data. More than 300 sequences, each more than 1500 elements in length, may be analyzed together. With the system scientists can classify elements, align sequences, edit them, find consensus patterns, and simultaneously generate oligomer frequency histograms and other statistics.

Expression and characterization of RNase III and Era proteins. Products of the rnc operon of Escherichia coli.

The synthesis rates of ribonuclease III (RNase III) and Era proteins are relatively low, and expression of the era gene is translationally coupled with expression of the rnc gene. Expression of both genes is negatively controlled by RNase III itself. We have constructed plasmids that overproduce RNase III and/or Era proteins under the control of the lambda PL promoter. A plasmid with the rnc gene under PL control expresses RNase III at levels greater than 40% of total cellular protein. Another plasmid with the era gene under PL control and a modified translation-initiation signal produces up to 80% of total cell protein as Era. Each protein has been purified using simple and rapid procedures. Purified RNase III protein specifically processes mRNA transcripts containing known RNase III sites. The purified Era protein binds GDP and GTP and has GTPase activity. Kinetic analysis shows that one molecule of GTP or GDP is bound/Era peptide with a Kd of 5.5 microM for GTP binding and 1.0 microM for GDP binding. The Km of the Era GTPase is 9.0 microM, and the maximum catalyzed rate of GTP hydrolyzed/min/mol of Era protein at 37 degrees C is 9.8 mmol.

Microplate assay of glutathione s-transferase activity for resistance detection in single-mosquito triturates.

1. Optimum conditions are described for a simple, rapid microplate assay that measures glutathione s-transferase (GST) activity accurately and precisely in small portions of single mosquito homogenates. 2. Up to 10 assay replicates were possible for individual adults and larvae. Concentration of GST activity in the head/thorax region allows blood-fed mosquitoes with abdomens removed to be used in assays. 3. The method allows the use of GST activity as a biochemical character in comparative studies of populations. 4. The microplate assay detects elevated GST activities associated with DDT resistance in Anopheles arabiensis.

Finger-loops, oncogenes, and metals. Claude Passmore Brown memorial lecture.

Certain DNA-binding proteins that regulate gene expression contain single or multiple copies of short polypeptide sequences, approximately 30 residues long, consisting of combinations of four Cys or His residues at defined spacing, so that Zn++ is complexed in tetrahedral coordination with the respective thiol-sulfur and/or imidazole-nitrogen atoms. The Zn++ ion evidently serves as a strut that stabilizes folding of the domain into a 'finger-loop', which is capable of site-specific binding to double-stranded DNA. This article reviews the evidence (a) that finger-loop domains have been highly conserved during evolution, (b) that they furnish one of the fundamental mechanisms for regulating gene expression, and (c) that a metal ion (e.g., Zn++) is required for binding of finger-loops to DNA and for their biological functions. The authors' search of amino acid sequences of 38 transforming proteins identified possible finger-loop domains in the myc, fms, fps, raf-1, rfp, src, syn, yes, erbA, int-1, and TGF-alpha gene-products. The search incidentally revealed possible finger-loop domains in human insulin receptor, which may provide a mechanistic explanation for recent observations that insulin, after binding to its cell surface receptor, is translocated to hepatocyte nuclei and becomes bound to chromatin. Zn++-coordination sites in finger-loop domains are proposed as potential targets for metal toxicity; substitution of Ni++, Co++, or Cd++ for Zn++ in finger-loops of transforming proteins is suggested as an hypothetical mechanism for metal carcinogenesis.