[Efficacy of Han-uvulopalatopharyngoplasty (HUPPP) combined with radiofrequency ablation of tongue base or HUPPP with traction of tongue base on moderate to severe patients with obstructive sleep apnea hypopnea syndrome (OSAHS):a multicenter randomized controlled trial].

Objective: To compare the therapeutic efficacy of Han-uvulopalatopharyngoplasty (HUPPP) combined with radiofrequency ablation of tongue base or HUPPP with traction of tongue base on moderate to severe patients with obstructive sleep apnea hypopnea syndrome (OSAHS). Methods: This is a multicenter randomized controlled trial. From March 2017 to July 2019, moderate to severe OSAHS patients from three clinical center in Shanghai who were intolerant to continuous positive airway pressure (CPAP) and with velopharyngeal and glossopharyngeal plane obstruction were enrolled in this study. According to the surgical type, they were 1∶1 randomized to HUPPP plus radiofrequency ablation of tongue base group (Ablation group) or HUPPP plus traction of tongue base group (Traction group). All patients completed over-night standard Polysomnography (PSG), upper-airway assessment (Friedman classification, Müller test, CT and cephalometric examination), preoperative routine examination, Epworth Sleepiness Scale (ESS) and Quebec sleep questionnaire (QSQ). Six to 12 months after operation, all the above-mentioned examinations were repeatedly performed. Changes of aforementioned variables before and after operation were assessed. Results: A total of 43 patients with moderate to severe OSAHS were enrolled in this study. One patient lost to follow-up, the remaining 21 were allocated to Ablation group and 21 were allocated to Traction group. The total therapeutic efficacy of all patients was 69.05% (61.90% in Ablation group and 76.19% in Traction group), but there was no statistical significance between the two groups (P= 0.317). The value of sleep scale score (ESS and QSQ), objective sleep variables (apnea-hypopnea index, oxygen saturation, percentage of time with blood oxygen less than 90% in total sleep time, oxygen desaturation index and micro-arousals) and upper airway cross-sectional area (palatopharyngeal and retrolingual area) of the two groups were improved (P<0.05), but the differences between the two groups were not statistically significant (P>0.05). Conclusion: For moderate to severe OSAHS who had glossopharyngeal plane obstruction, both HUPPP plus radiofrequency ablation of tongue base or HUPPP plus traction of tongue base are effective treatment for OSAHS, and the curative effect is similar. The choice of surgical type could be selected according to patient's or surgical conditions.

Factor structure of 12 items in the Chinese Health Questionnaire among the elderly population in mainland China.

OBJECTIVES: The 12-item Chinese Health Questionnaire (CHQ-12) has been widely used for screening mental disorders. This study aims to examine the internal consistency and factor structure of CHQ-12 and its suitability for use in the elderly Chinese population. STUDY DESIGN: This is a cross-sectional study. METHODS: A total of 8526 elderly people aged ≥60 years from 11 cities in Shanxi Province were selected for participation in this study by stratified random cluster sampling. Cronbach's alpha was employed to assess internal consistency. An exploratory factor analysis (EFA) was performed to explore the underlying factor structure of the CHQ-12 in the elderly. A confirmatory factor analysis (CFA) was then conducted to test and compare the goodness-of-fit between possible factor structure obtained from the EFA and the unidimensional structure, which was originally recommended. RESULTS: The Cronbach's alpha for CHQ-12 was 0.838. The EFA extracted three factors, which explained 55.985% of the total variance of the data. The CFA of the three-factor model resulted in an acceptable model fit (Comparative Fit Index = 0.98, Tucker-Lewis Index = 0.97, Normed Fit Index = 0.98, Expected Cross-Validation Index = 0.28, Root-Mean-Square Error of Approximation = 0.071). The item loadings ranged from 0.58 to 0.82. Correlation coefficients among the three factors ranged from 0.40 to 0.75. CONCLUSIONS: The CHQ-12 presented satisfactory internal consistency and structural validity in the Chinese elderly population. The CFA of the three-factor structure expressed a preferred model fit in comparison to the unidimensional model. The three-factor structure of the CHQ-12 interpreted three different aspects of mental health: somatic symptoms, anxiety and worry, and depression/poor family relationship.

[The role of EBUS-TBNA in the systematic evaluation of lymph node staging and resectability analysis in non-small cell lung cancer].

Objective: To evaluate the role of endobronchial ultrasound guided transbronchial needle aspiration (EBUS-TBNA) in lymph node staging and resectability assessment of patients with non-small cell lung cancer (NSCLC). Methods: The clinical data of 154 patients with NSCLC who underwent EBUS-TBNA from March 2015 to December 2018 were collected. All accessible mediastinal and hilar lymph nodes were systematically explored and punctured using EBUS-TBNA. EBUS-TBNA and CT were used for preoperative staging and resectability evaluation. Results: The sensitivity, specificity and accuracy of EBUS-TBNA were 94.2%, 100.0% and 96.0%, respectively, while those of CT were 89.9%, 31.8% and 72.0%, respectively. The differences were statistically significant (P<0.05). The sensitivity, specificity and accuracy of EBUS-TBNA in lymph nodes with short diameter less than 15 mm were 92.4%, 100.0% and 96.0%, respectively, while those of CT were 80.7%, 34.8% and 60.1%, respectively, with statistical differences (P<0.05). The staging of 62 patients was changed, 27 cases were up-regulated and 35 cases were down-regulated. Among them, 32 cases had been changed to resectable. The evaluating resectability of EBUS-TBNA showed excellent consistency with that of pathological results (Kappa=0.95). The sensitivity and specificity were 100.0% and 97.2%, respectively. Conclusion: EBUS-TBNA can systemically evaluate the metastatic status of NSCLC patients and improve the accuracy of preoperative lymph node staging and resectability assessment.

[The research of sentinel lymph node biopsy after neoadjuvant chemotherapy for breast cancer patients].

With the extensive use of sentinel lymph node biopsy (SLNB), some breast cancer patients could avoid axillary lymph node dissection (ALND) and its complications. Neoadjuvant chemotherapy plays an important role in the multimodality therapies of breast cancer. After neoadjuvant chemotherapy, some patients with breast cancer were down-staged from positive axillary lymph node (cN+ ) to clinically negative (cN0). For these patients, the feasibility and safety of sentinel lymph node biopsy remains controversial. However, with the application of new technologies, SLNB is expected to become the main treatment for breast cancer patients with stage cN0 after neoadjuvant chemotherapy.

[Relationship between smoking and the severity of OSA].

Objective:To explore the relationship between smoking and the severity of OSA. Method:There were 719 patients included in the study, who were accompanied by snoring, daytime sleepiness and other symptoms. Laboratory-based polysomnographic variables （including AHI, oxygen desaturation index and microarousal index, etc.）, and anthropometric measurements （including weight, neck circumference, waist circumference, hip circumference etc.） were collected for all participants. The severity of OSA was determined by AHI. No OSA was defined as AHI<5, mild OSA as AHI of 5 to 15，moderate OSA as AHI of >15 to 30, and severe OSA as AHI of >30. Smoking severity was determined by the smoking index （SI）. Light smoke was defined as SI<200, moderate smoke was as SI 200 to 400, and severe smoke as SI>400. Result:There were 138 cases of non-OSA and 581 cases of OSA. There were 381 non-smokers, 279 smokers and 59 quit smokers. The smoking rate of OSA group was significantly higher than that of non-OSA group （41.5% vs. 27.5%,P<0.01）. After excluding 59 quit smokers, the remaining 660 subjects were divided into four groups according to the severity of smoking, then each group was further divided into four groups according to OSA severity. Unadjusted analysis showed that OSA severity positively correlated with smoking severity （r=0.203,P<0.01）. The positive correlation remained significant after further adjustment for age, BMI and waist-hip ratio. In addition, logistic regression analysis showed that compared to non-smokers, the odd ratios for OSA in moderate smokers were 1.72 （95%CI 1.08-2.75） and in severe smokers were 2.68 （95%CI 1.61-4.46）, after adjustment for age, BMI and waist-hip ratio. Conclusion:The severity of smoking significantly correlated with the severity of OSA. There was increased risk of OSA in patients with severe smoke. The correlation was independent of some confounders such as age and obesity.

MiR-216a-5p act as a tumor suppressor, regulating the cell proliferation and metastasis by targeting PAK2 in breast cancer.

OBJECTIVE: Our study aimed to investigate the expression of microRNA-216a-5p (miR-216a-5p) in breast cancer (BC) and its effect on the proliferation and metastasis of BC cells by regulating the expression of p21-activated protein kinase 2 (PAK2) gene. PATIENTS AND METHODS: A total of 50 cases of cancer tissue specimens and corresponding para-carcinoma normal tissue specimens were collected from the breast surgery department of our hospital from July 2016 to December 2017. BC MCF-7 cell line and normal breast epithelial MCF-10A cells were cultured. MiR-NC (negative control), LV-p21-activated protein kinase 2 (PAK2) and/or miR-216a-5p mimics were synthesized and transfected. The protein and mRNA expression level in BC tissues and cells were detected by Western blot and quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) assay, respectively. Additionally, the Luciferase Reporter Assays, cell proliferation detection, clone formation assays and transwell migration and invasion assay were performed to determine the functional alteration of BC cells, respectively. RESULTS: The results of qRT-PCR demonstrated that miR-216a-5p was decreased in both BC tissues and cells compared with that in normal controls. Online target gene prediction software and Dual-Luciferase reporter assay were used for target identification, and PAK2 was identified as a functional target of miR-216a-5p in BC cells. The results were further clarified with the Western blot (WB) experiment. In vitro, cell functions were detected by Cell Counting Kit-8 (CCK-8), crystal violet staining and transwell experiment, respectively. The results indicated that decreased expression of PAK2 resulting from the up-regulation of miR-216a-5p could restrain the proliferation, clone formation, invasion and migration abilities of BC cells. CONCLUSIONS: We showed that miR-216a-5p played a role as antioncogene in BC, which provides a new therapeutic target for the treatment of BC.

Binding site differences revealed by crystal structures of Plasmodium falciparum and bovine acyl-CoA binding protein.

Acyl-CoA binding protein (ACBP) maintains a pool of fatty acyl-CoA molecules in the cell and plays a role in fatty acid metabolism. The biochemical properties of Plasmodium falciparum ACBP are described together with the 2.0 A resolution crystal structures of a P. falciparum ACBP-acyl-CoA complex and of bovine ACBP in two crystal forms. Overall, the bovine ACBP crystal structures are similar to the NMR structures published previously; however, the bovine and parasite ACBP structures are less similar. The parasite ACBP is shown to have a different ligand-binding pocket, leading to an acyl-CoA binding specificity different from that of bovine ACBP. Several non-conservative differences in residues that interact with the ligand were identified between the mammalian and parasite ACBPs. These, together with measured binding-specificity differences, suggest that there is a potential for the design of molecules that might selectively block the acyl-CoA binding site.

Ethanol pre-exposure suppresses HIV-1 glycoprotein 120-induced neuronal degeneration by abrogating endogenous glutamate/Ca2+-mediated neurotoxicity.

The neurotoxic mechanism of HIV-1 envelope glycoprotein 120 (gp120) involves glutamatergic (NMDA) receptor/Ca2+-dependent excitotoxicity, mediated in part via glia. Pro-inflammatory cytokines also may have roles. We have reported that pre-exposure of brain cultures to 'physiological' ethanol concentrations (20-30 mM) protects against neuronal damage from HIV-1 gp120, but not from the direct receptor agonist, NMDA. Using lactate dehydrogenase assays and propidium iodide staining of rat organotypic hippocampal-entorhinal cortical slice cultures we determined that ethanol's suppression of gp120 neurotoxicity required at least 4 days of pretreatment. The gp120-induced neurotoxicity was accompanied by interleukin-6 elevations that were not affected by the pretreatment. However, gp120 induced substantial, early increases in extracellular glutamate levels that were blocked by ethanol pretreatment, conceivably abrogating excitotoxicity. Consistent with abrogation of excitotoxic pathways, fura-2 imaging showed selective deficits in gp120-dependent intracellular Ca2+ responses in ethanol-pretreated slices. Gp120 is believed to increase glutamate levels by both stimulating release and inhibiting (re)uptake. Results with a labeled glutamate analog, D-[3H]aspartate, revealed that gp120's inhibition of glutamate uptake, rather than its stimulation of release, was abolished after ethanol. Further studies indicated that two converging effects of ethanol pretreatment may underlie the abolishment of gp120-mediated glutamate uptake inhibition: (a) blockade of gp120-induced release (ostensibly from glia) of arachidonic acid, an inhibitor of astroglial glutamate reuptake, and (b) modest proliferation and activation of astroglia upon gp120 stimulation--which are likely to augment glutamate transporters. Thus, as with gp120 itself, glia and glutamate/arachidonic acid regulation appear to be important targets for ethanol. Since moderate ethanol consumption is as common among HIV-infected individuals as in the general population, this newly recognized neuroprotective (and apparently anti-excitotoxic) effect of ethanol withdrawal in vitro could be important, but it requires further study before its significance, if any, is understood.

HIV-I gpI20 neurotoxicity in brain cultures is prevented by moderate ethanol pretreatment.

The HIV-1 coat protein gp 20, a potent neurotoxin that may underlie AIDS dementia, activates glia to cause neurotoxicity via the NMDA receptor and perhaps other routes. We find that pretreating cultures of rat organotypic cortical/hippocampal slices or cerebellar granule cells subchronically with ethanol in physiological concentrations (20-30 mM; 6 days) largely or even completely inhibits neurodegeneration due to gp120. However, NMDA-induced neurotoxicity appears unaffected by moderate ethanol pretreatment, indicating that ethanol's neuroprotection against gp120 is upstream of the NMDA receptor, possibly at a glial activation stage. The results could lead to a better understanding of relationships between ethanol, glia and neurodegeneration, particularly in AIDS.

Crystallographic evidence for substrate ring distortion and protein conformational changes during catalysis in cellobiohydrolase Ce16A from trichoderma reesei.

BACKGROUND: Cel6A is one of the two cellobiohydrolases produced by Trichoderma reesei. The catalytic core has a structure that is a variation of the classic TIM barrel. The active site is located inside a tunnel, the roof of which is formed mainly by a pair of loops. RESULTS: We describe three new ligand complexes. One is the structure of the wild-type enzyme in complex with a nonhydrolysable cello-oligosaccharide, methyl 4-S-beta-cellobiosyl-4-thio-beta-cellobioside (Glc)(2)-S-(Glc)(2), which differs from a cellotetraose in the nature of the central glycosidic linkage where a sulphur atom replaces an oxygen atom. The second structure is a mutant, Y169F, in complex with the same ligand, and the third is the wild-type enzyme in complex with m-iodobenzyl beta-D-glucopyranosyl-beta(1,4)-D-xylopyranoside (IBXG). CONCLUSIONS: The (Glc)(2)-S-(Glc)(2) ligand binds in the -2 to +2 sites in both the wild-type and mutant enzymes. The glucosyl unit in the -1 site is distorted from the usual chair conformation in both structures. The IBXG ligand binds in the -2 to +1 sites, with the xylosyl unit in the -1 site where it adopts the energetically favourable chair conformation. The -1 site glucosyl of the (Glc)(2)-S-(Glc)(2) ligand is unable to take on this conformation because of steric clashes with the protein. The crystallographic results show that one of the tunnel-forming loops in Cel6A is sensitive to modifications at the active site, and is able to take on a number of different conformations. One of the conformational changes disrupts a set of interactions at the active site that we propose is an integral part of the reaction mechanism.

Brain damage due to episodic alcohol exposure in vivo and in vitro: furosemide neuroprotection implicates edema-based mechanism.

Adult rats intubated with a single dose of ethanol (alcohol; approximately 5 g/kg) for 5 to 10 successive days incur neurodegeneration in the entorhinal cortex, dentate gyrus, and olfactory bulbs accompanied by cerebrocortical edema and electrolyte (Na+, K+) accumulation. The brain damage is not lessened by cotreatment with the NMDA receptor antagonist MK-801; also, as reported elsewhere, MK-801 as well as non-NMDA receptor and Ca2+ channel antagonists are not neuroprotective in a similar, but more compressed, intoxication protocol. However, cotreatment with the electrolyte transport inhibitor/diuretic furosemide reduces alcohol-dependent cerebrocortical damage by 75-85% while preventing brain hydration and electrolyte elevations; olfactory bulb neurodegeneration is not attenuated. In parallel in vitro studies, rat organotypic entorhinal/hippocampal slice cultures exposed to alcohol (50-200 mM) 15 h/day for 6 days, mirroring episodic intoxication in vivo, demonstrate concentration-related release of the cytotoxic indicator, lactate dehydrogenase. Analogous to the in vivo findings, furosemide blocks this alcohol-induced in vitro cytotoxicity. Our results showing neuroprotection by furosemide indicate that brain edema and swelling are essential events in the brain damage induced by episodic alcohol exposure. Furosemide and related agents might be useful as neuroprotective agents in alcohol abuse. We suggest that the neurodegeneration is elicited in part by edema-dependent oxidative stress, but the regional selectivity of the damage may be best explained by physical (mechanical) compression of the limbic cortex against the adjacent tympanic bulla and subsequent neuronal cytoskeletal collapse. A scheme for these apparently nonexcitotoxic metabolic and mechanical pathways initiated by repeated alcohol exposure is proposed.

The crystal structure of the catalytic core domain of endoglucanase I from Trichoderma reesei at 3.6 A resolution, and a comparison with related enzymes.

Cellulose is the most abundant polymer in the biosphere. Although generally resistant to degradation, it may be hydrolysed by cellulolytic organisms that have evolved a variety of structurally distinct enzymes, cellobiohydrolases and endoglucanases, for this purpose. Endoglucanase I (EG I) is the major endoglucanase produced by the cellulolytic fungus Trichoderma reesei, accounting for 5 to 10% of the total amount of cellulases produced by this organism. Together with EG I from Humicola insolens and T. reesei cellobiohydrolase I (CBH I), the enzyme is classified into family 7 of the glycosyl hydrolases, and it catalyses hydrolysis with a net retention of the anomeric configuration. The structure of the catalytic core domain (residues 1 to 371) of EG I from T. reesei has been determined at 3.6 A resolution by the molecular replacement method using the structures of T. reesei CBH I and H. insolens EG I as search models. By employing the 2-fold non-crystallographic symmetry (NCS), the structure was refined successfully, despite the limited resolution. The final model has an R-factor of 0.201 (Rfree 0.258). The structure of EG I reveals an extended, open substrate-binding cleft, rather than a tunnel as found in the homologous cellobiohydrolase CBH I. This confirms the earlier proposal that the tunnel-forming loops in CBH I have been deleted in EG I, which has resulted in an open active site in EG I, enabling it to function as an endoglucanase. Comparison of the structure of EG I with several related enzymes reveals structural similarities, and differences that relate to their biological function in degrading particular substrates. A possible structural explanation of the drastically different pH profiles of T. reesei and H. insolens EG I is proposed.

Binge ethanol-induced brain damage in rats: effect of inhibitors of nitric oxide synthase.

Testing the possible role of endogenous nitric oxide (NO) in the neurotoxicity of ethanol, we examined how two different NO synthase (NOS) inhibitors affected the extent cerebrocortical and olfactory neuronal damage in a modified "binge intoxication" rat model (Collins et al., Alcohol Clin. Exp. Res. 20:284-292, 1996). Male rats intragastrically fed ethanol (6.5 to 12 g/kg/day) in nutrient solution three times daily for 4 days also received NG-nitro-L-arginine methyl ester by chronic intracerebroventricular infusion or 7-nitro-indazole by daily intraperitoneal injection; control rats were given nutrient solution only and/or vehicles. Blood ethanol levels did not differ among the ethanol-treated groups. The amount of ethanol-dependent neuronal degeneration in the entorihinal cortex, dentate gyrus, and olfactory bulb glomeruli--visualized with the de Olmos cupric silver stain and quantitatively assessed in the binge-intoxicated rats--was either unchanged or significantly increased by the NOS inhibitors. Although the efficacies of the inhibitors cannot be directly compared because of various NOS forms were probably inhibited to differing extents, the results do not support the idea that endogenous NO is a neurotoxic mediator of ethanol's effects. Rather NO may have a modest neuroprotectant role in this model of early brain damage induced by ethanol. In addition, the NOS that is localized histochemically as NADPH diaphorase was present primarily in regions and/or cells not damaged by binge ethanol treatment. Assuming that NADPH diaphorase represents most of the NO forming enzyme(s) this suggests a transcellular mechanism for NO. A further observation was that hippocampal CA pyramidal neuron degeneration was extensive in rats infused centrally with NG-nitro-L-arginine methyl ester.

Towards the automatic interpretation of macromolecular electron-density maps: qualitative and quantitative matching of protein sequence to map.

The matching of the known polypeptide sequence to the electron density is a critical step in solving protein structures by the crystallographic method. Tools have been developed to help in defining the placement of the sequence, both qualitatively and quantitatively. They have been tested with good results on two proteins whose structures were solved by the MIR method.

Continuous exposure of cultured rat cerebellar macroneurons to ethanol-depressed NMDA and KCl-stimulated elevations of intracellular calcium.

This series of experiments measured ethanol-induced changes in levels of free intracellular calcium. Cerebellar macroneurons, harvested from rat embryos on embryonic day 17, were cultured in the presence of 75 mM ethanol for 24, 48, or 96 hr. Intracellular calcium concentrations in control and ethanol-exposed neurons did not differ after 24 hr, but they were significantly elevated in the neurons exposed to ethanol for 48 or 96 hr. Similarly, increases in intracellular calcium elicited by stimulation with 50 microM NMDA were not significantly different in control and ethanol-exposed neurons after 24 hr. After 48 and 96 hr, however, NMDA-stimulated increases in intracellular calcium levels in control neurons were significantly greater than in the ethanol-exposed neurons. These results showed that, when calcium levels were elevated by prolonged exposure to ethanol, the neurons were significantly less responsive to NMDA stimulation. Increases in intracellular calcium elicited by stimulation with 30 mM KCI were not significantly different in the control and treated neurons after 24 and 48 hr of ethanol exposure. After 96 hr of exposure to ethanol, however, there was a significant increase in intracellular calcium levels in control neurons following KCI stimulation, but not in the ethanol-exposed neurons. The fact that neuronal responses to KCI stimulation were depressed only following 96 hr of exposure to ethanol makes it unlikely that voltage-regulated channels were the primary mediators of the ethanol-induced elevations in intracellular calcium in chronically exposed neurons.

The effects of target tissues on the outgrowth of chick cutaneous and muscle sensory neurons.

In some developing systems, growing axons are attracted to their target site by a diffusible molecule released by the target tissue. In the chick hindlimb, this mechanism could explain how axons, after having reached the plexus region, grow to muscle or to skin. To begin to test this possibility for limb-innervating sensory neurons, we cocultured dorsal root ganglion explants and potential target tissues in three-dimensional collagen gels. In particular, we wanted to know if target tissues, at early stages of development, specifically attract axons of only the appropriate type of sensory neuron. To identify each type of sensory neuron, we used DiI to retrogradely label either cutaneous or muscle sensory neurons in the embryo, prior to culturing. The results showed that dermal and muscle explants could each enhance the outgrowth of both cutaneous and muscle sensory axons. In contrast, the epidermis and the connective tissue associated with developing muscle appeared to inhibit the outgrowth of both cutaneous and muscle sensory axons. These results suggest that, in the embryo, the dermis and muscle cells both release diffusible factors that cause sensory axons to diverge from the plexus, extend toward the sources of these factors, and thereby form discrete peripheral nerves. The inhibitory effects of epidermis and muscle-associated connective tissue may serve to limit the growth of sensory axons to the structures, i.e., dermis and muscle cells, that ultimately receive sensory innervation. However, since for each of the difference types of limb tissue, the responses of cutaneous and muscle sensory neurons were always similar to one another, sensory axons must not be responding to target-derived factors when they decide whether to grow to skin or to muscle.

[Production and identification of a monoclonal antibody against a tumor associated antigen from ovarian epithelial carcinoma].

The splenocyte of BALB/c mice, immunized with soluble and ovarian serous papillary cystedenoma-associated antigen CA925, was fused with NS-1 myeloma cell strain to produce a monoclonal antibody. 5 cell strains of the monoclonal antibody with continuous secretion were obtained. The fusion openings were used as code names: OC4D9, OC7E10, OC1B4, OC3B8, and OC9B9, their titers were 1: 10(7), 1: 10(7), 1: 10(6), 1: 10(4) and 1: 10(5) respectively. Ig subgroups were IgM and IgG. The chromosomes showed the characteristics of their parents. The 5 strains of the monoclonal antibody obtained presented specific reactions to CA925, no positive reaction with normal ovarian tissues or leiomyoma of the uterus were seen, and there were weak reactions with carcinomas of the lung, the stomach, the kidney, the esophagus and the rectum. Immunohistochemical tests proved that the monoclonal antibody produced was specific against ovarian epithelial carcinoma, and it might offer a means to the diagnosis and treatment of malignant ovarian tumors.

An evaluation of the use of databases in protein structure refinement.

The speed of electron-density fitting during X-ray structure solution and refinement, and the quality of the protein model resulting, can both be enhanced by the use of databases of main- and side-chain conformations. Three structures are compared in this report, one refined at high resolution (1.7 A), and two at lower resolutions using either the database method (2.4 A resolution) or more traditional empirical electron-density fitting (1.9 A resolution). An analysis of peptide orientation was used as an aid in finding unusual portions of main-chain structure. The fit of side chains to known rotamer conformations was used to help determine the accuracy of these atomic positions. In addition, the use of an objective measure of the fit of structures to electron-density maps was evaluated, both alone and in combination with side-chain conformational information.

The 1.7 A refined X-ray structure of the periplasmic glucose/galactose receptor from Salmonella typhimurium.

The X-ray structure of the periplasmic glucose/galactose receptor (binding protein) of Salmonella typhimurium (GBP-S) has been refined at 1.7 A resolution with an R-factor of 19.0%. The model contains all 309 residues of the amino acid sequence, 153 water molecules, a calcium ion and beta-D-galactose. The protein consists of two very similar structural domains, each of which is composed a core of parallel beta-sheet flanked on both sides by alpha-helices. Three short stretches of amino acid chain (from symmetrically related portions of the structure) link the domains, and presumably act as a hinge to allow their relative movement in functionally important conformational changes. Galactose is bound between the domains, interacting with a number of side-chains from the loops lining the binding cleft. A combination of hydrogen bonding, hydrophobic and steric effects give rise to tight binding (dissociation constant 0.2 microM) and high specificity. Of nine hydrogen bonding groups, three are aspartate, three asparagine, one histidine (unprotonated), one arginine and one water, contributing 13 hydrogen bonds in total. Additional residues pack against (primarily) non-polar faces of the sugar molecule. The precise arrangement of the hydrogen bonding and hydrophobic residues results in an enclosed binding site with a shape that is a composite of those of the allowed sugar molecules. It is presumed that ligands bind to a more open form of the receptor that then closes by rotation in the hinge. Comparison with the GBP-S structure solved earlier in complex with glucose shows no significant changes, even for the aspartate residue most directly involved with the different sugars. Comparison with the galactose/glucose receptor of Escherichia coli indicates that these two proteins are very similar in overall structure, with the main difference being a 2 to 3 degrees rotation in the hinge. This difference appears to be the result of different crystal packing for the two proteins; it is likely that both conformations are normally found in solution.