Solution structure and function of a conserved protein SP14.3 encoded by an essential Streptococcus pneumoniae gene.

Streptococcus pneumoniae is a major human pathogen that causes high mortality and morbidity rates and has developed resistance to many antibiotics. The genome of S. pneumoniae has recently been completely sequenced revealing many genes encoding hypothetical proteins of unknown function. We have found that the gene encoding one such conserved protein, SP14.3, is essential for growth of S. pneumonia. Since it is essential, SP14.3 represents a potential target for drug discovery. Here, we describe the three-dimensional solution structure of SP14.3 as determined by NMR spectroscopy. The structure consists of two domains each with an alpha/beta-fold. The N-terminal domain contains two alpha-helices and a three-stranded beta-sheet, while the C-terminal domain is composed of one alpha-helix and a five-stranded beta-sheet. The N-terminal domain of the protein contains a highly negatively charged surface and resembles the fold of the N-terminal domain of Thermus thermophilus ribosomal protein S3. The C-terminal domain has a protein fold similar to human small nuclear ribonucleoprotein Sm D3 and Haloarcula marismortui ribosomal protein L21E. The two domains of the protein tumble in solution overall as a whole with an overall molecular rotational correlation time (tau(m)) of 12.9 ns at 25 degrees C. The relative orientation of the two domains is not defined by the nuclear Overhauser effect data. Indeed, residual dipolar couplings and the structure calculations indicate that the relative orientation of the two domains is not rigidly oriented with respect to one another in solution.

"Love's labours lost": failure to implement mass vaccination against group A meningococcal meningitis in sub-Saharan Africa.

PIP: Despite the availability of a safe, effective polysaccharide vaccine, group A meningococcal meningitis epidemics persist in sub-Saharan Africa. In October 1996, there were almost 150,000 reported cases and 15,000 deaths, the majority of which involved children. At 3 months of age, induction of protective group A meningococcal antibody levels requires 2 injections at least 1 month apart. Reinjection of 5-year-old children increases group A antibodies to long-term protective levels. During meningitis epidemics in Nigeria, Mali, and Rwanda, fatality was significantly reduced in areas where scarce vaccine was administered selectively. Although effective on an individual basis, selective vaccination is unable to control meningitis epidemics. In Chad, mass vaccination of the entire population (excluding infants under 12 months) eliminated the disease. Successful mass vaccination against group A meningococcal epidemics also has been reported in Saudi Arabia, China, and refugee camps in Africa. Although cost is cited as an obstacle to routine mass vaccination to prevent meningococcal meningitis in South Africa, prevention is the least expensive approach to disease control. It is recommended that the entire population of Africa's meningitis belt receive group A meningococcal vaccine in accordance with the recommended age schedule in a mass vaccination program.^ieng

The forty-day rest period and infant feeding practices among Negev Bedouin Arab women in Israel.

Four hundred and twelve Bedouin Arab women were interviewed within 48 hours of delivering a newborn in 1982 and two months later to examine the effect of social support during the forty-day postpartum rest period on the incidence of exclusive breast feeding at two months. Whereas the distribution of infant feeding practices at two months did not differ among women with and without support during the "forty days," the number of days of help during the "forty days" was significantly associated with type of feeding at two months. The longer the duration of help, the greater were the odds of maintaining exclusive breast feeding after adjustment for covariates by multinomial logistic regression analysis. The association between duration of help and infant feeding practices at two months was modified by the interview month and the feeding practices of the respondents' primary social supports. More respondents with infants who reached two months of age in May and June were exclusively breast feeding than respondents with infants who reached two months in the hot, dry months of July and August. Also, respondents with sisters-in-law who breast fed alone or in combination with the bottle were feeding their infants in the same way after adjustment for covariates. Given the beneficial effects of breast feeding on infant health and development, these findings identify potential avenues of intervention for programs promoting breast feeding.

PIP: In March-June 1982, female Arab students at Ben Gurion University in Beer Sheva, Israel, interviewed 412 mothers within 2 days after they delivered healthy newborns either at the Soroka Medical Center or at home to examine the link between family support during the traditional 40-day rest period and the incidence of exclusive breast feeding during the infant's first 2 months of life. Mothers exclusively breast fed only 86% of the newborns. By 2 months, this percentage fell to just 24%. 22% did not receive any help with chores during the 40-day rest period and only 59% of those who did receive such help had this help for 40 days. These effects were probably a result of the transition from the traditional seminomadic to a sedentary life style. Even though the distribution of infant feeding practices between those who received help with chores during the 40-day rest period and those who did not were not greatly different, the duration of help with cooking did differ between the 2 groups (p = .02). 31% of mothers who had someone help with cooking for =or 40 days exclusively breast fed while only 24% of mothers who had someone help with cooking for 40 days did so. Mothers whose infants were 2 months old in the hot and dry months of July or August were 5 times more likely to switch from breast feeding to breast and bottle feeding than were those whose infants were 2 months old in May or June (p = .006). They were also 4 times more likely to switch from breast feeding to feeding their infants cereal, breast milk and liquids from a bottle (p = .005). Mothers tended to follow the same infant feeding practices as their sisters-in-laws (p = .06 and .01 respectively). The reduction of help with chores during the first 40 days postpartum may adversely affect the health and development of Bedouin mothers and their children.

Association of fertility, fitness and longevity with the murine Ah locus among (C57BL/6N) (C3H/HeN) recombinant inbred lines.

The Ah locus encodes a cytosolic receptor which controls the induction of enzymes that metabolize drugs, chemical carcinogens, and other environmental pollutants. B6NXC3N recombinant inbred lines have been developed from the progenitors C57BL/6N and C3H/HeN inbred mouse strains. Ah phenotyping at each generation has resulted in the establishment of some lines containing high levels of the high-affinity Ah receptor; other lines, very low levels. A genetic model involving two unlinked loci is offered to explain the distribution of Ah receptor levels among (C57BL/6N) (C3H/HeN)F2 individuals. Between generations 7 and 13, individual females and males from the B6NXC3N recombinant inbred lines were crossed with DBA/2N males and females. Presence of high levels of the high-affinity Ah receptor in both female and male B6NXC3N mice was found to be associated with greater fertility, fitness, and longer life span. The data suggest that these parameters are correlated with the Ah locus or a closely segregating gene.

Pantetheinase activity and cysteamine content in cystinotic and normal fibroblasts and leukocytes.

Cysteamine is the most effective agent known for the reduction of the elevated cystine content of cells from patients with cystinosis. A defect in endogenous cysteamine generation could account for many of the metabolic features of this disorder. To test this hypothesis, we have developed improved methods for measuring pantetheinase (cysteamine-generating) activity and intracellular cysteamine levels and used these methods to measure such parameters in cystinotic and normal leukocytes and cultured skin fibroblasts. Pantetheinase activity as defined in the test was similar in extracts of cystinotic and normal cells [leucocytes, normal, 78 +/- 15 (S.E.), cystinotic, 56+/- 6.4; fibroblasts, normal, 9.4 +/- 1.5; cystinotic, 7.7 +/- 1.7]. Cysteamine levels were normal in leukocytes from cystinotics receiving no cysteamine or doses of oral cysteamine too low to reduce leukocyte cystine content. The results indicate that the cause of cystinosis is unlikely to be related to a failure to generate of sustain normal intracellular cysteamine levels.

Glycine and benzoate conjugation and glycine acyltransferase activity in the developing and adult rat: possible relationships to nonketotic hyperglycinemia.

We report investigations of benzoate and glycine metabolism and glycine acyltransferase activity in rats. These studies provide insights related to the therapy and pathophysiology of human nonketotic hyperglycinemia. Liver acyltransferase activity increased sharply postnatally from low levels at birth, but transferase activity was absent in the brain. The enzyme level was unchanged in either organ after administration of 2,3,7,8-tetrachlordibenzodioxin, phenobarbital, benzoate or a high glycine diet. Brain and liver glycine levels remained unaltered during acute or chronic benzoate-induced reductions in plasma glycine levels. Plasma and brain glycine contents were measured in rats at different ages following a single injection of 3 mg glycine/g body weight; after injection, glycine levels in the brain were comparable in severely symptomatic neonatal rats and older asymptomatic rats, suggesting a similar glycine influx but a selective susceptibility of the newborn brain to toxicity from acute hyperglycinemia. When a 3.4% glycine diet was ingested for up to 30 days ad libitum, levels of plasma glycine rose about 4- to 5-fold from those achieved on a diet containing one tenth as much glycine, but brain and liver glycine concentrations increased only 2-fold or less in the chronically hyperglycinemic animals.

Time estimation and hyperactivity, replication.

Three groups of boys included 135 of normal IQ, showing normal activity; 6 were hyperactive and of normal IQ, and 6 showed normal activity but low IQ. Their ability to estimate a 30-sec. interval showed only the 6 normally active boys of low IQ had significantly different elapsed and estimated times. Time estimation is not clinically useful for identifying hyperactive boys.

A model of fulminant hepatic failure in the rabbit.

A highly reproducible model of fulminant hepatic failure was developed by administering intravenously the selective hepatotoxin galactosamine hydrochloride (4.25 mmoles per kg) to genetically uniform rabbits. The great majority of rabbits died between 21 and 44 hr after injection following a period of coma which lasted 2.6 hr on average. Serum biochemical tests and liver histology reflected massive hepatic injury. Changes in plasma ammonia and amino acid concentrations, in coagulation parameters, and in the electroencephalogram were similar to those found in human fulminant hepatic failure. This model appears promising for future studies of the pathogenesis and treatment of fulminant hepatic failure.

Proteolytic fragmentation of fibrinogen. II. kinetic modeling of the digestion of human and bovine fibrinogen plasmin or trypsin.

The kinetic data presented in the previous paper (Mihalyi, E., et al. (1976), Biochemistry 15, preceding paper in this issue), with respect to the fragmentation of human the bovine fibrinogen by either plasmin or trypsin, were compared with several chemical kinetic models. The models were derived mathematically on the basis of the three-nodular structure of fibrinogen (Hall, C.E., and Sayter, H.S. (1959), J. BiophysBiochem. Ctyol. 5, 11) and the asymmetrical cleavage sequence first proposed by Marder, V.J., et. al. ((1969) J. Biol. Chem. 244, 2111). The parameters were determined by nonlinear curve fitting. The whole process could be described accurately by only two rate constants. Several variant models were tested and, although a clear cut choice cannot be made, one of these, the protected three-bonds model, appears to give the best fit in most cases. This model assumes that the chain segment that distinguishes F from X protects certain other chains (the bonds) from proteolytic cleavage.

Redox properties of beta-type cytochromes in Escherichia coli and rat liver mitochondria and techniques for their analysis.

We describe here apparatus and procedures for conducting potentiometric titrations and for analyzing the collected data in terms of the number of components present, their amounts and their midpoint potentials. Using these procedures we have determined the presence of three forms of cytochrome b1 in Escherichia coli with midpoint potentials at pH 7.1 OF about minus 50, plus 110 and plus 220 mV. We were not able to demonstrate a change in any of these potentials by the addition of phosphate, ATP, or 2, 4-dinitrophenol. We have been able to confirm the presence of two forms of cytochrome b in non-energized mitochondria and the apparent conversion of the low-potential component to a new high potential component upon energization of the mitochondria. However we cite further experimental data that question the actual conversion of one form of cytochrome b to another. An alternative interpretation based on our analysis suggests that the high voltage component may be present in a masked form in the non-energized mitochondria.