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Biomedical subjects

X C Zhang

Publications and source records attributed to X C Zhang.

At least 19 recordsLinked to original sources
PubMed2000

Structure of the protease domain of memapsin 2 (beta-secretase) complexed with inhibitor.

Memapsin 2 (beta-secretase) is a membrane-associated aspartic protease involved in the production of beta-amyloid peptide in Alzheimer's disease and is a major target for drug design. We determined the crystal structure of the protease domain of human memapsin 2 complexed to an eight-residue inhibitor at 1.9 angstrom resolution. The active site of memapsin 2 is more open and less hydrophobic than that of other human aspartic proteases. The subsite locations from S4 to S2' are well defined. A kink of the inhibitor chain at P2' and the change of chain direction of P3' and P4' may be mimicked to provide inhibitor selectivity.

Amyloid Precursor Protein Secretases↗
PubMed2000

Human plasminogen catalytic domain undergoes an unusual conformational change upon activation.

Activation of the serine protease plasmin from its zymogen, plasminogen, is the key step in fibrinolysis leading to blood clot dissolution. It also plays critical roles in cell migration, such as in tumor metastasis. Here, we report the crystal structure of an inactive S741A mutant of human plasminogen catalytic domain at 2.0 A resolution. This structure permits a direct comparison with that of the plasmin catalytic unit. Unique conformational differences are present between these two structures that are not seen in other zymogen-enzyme pairs of the trypsin family. The functional significance of these differences and the structural basis of plasminogen activation is discussed in the light of this new structure.

Amino Acid Sequence↗
PubMed2000

Molecular mechanisms of plasminogen activation: bacterial cofactors provide clues.

Plasminogen activation is a key event in the fibrinolytic system that results in the dissolution of blood clots, and also promotes cell migration and tissue remodelling. The recent structure determinations of microplasmin in complex with the bacterial plasminogen activators staphylokinase and streptokinase have provided novel insights into the molecular mechanisms of plasminogen activation and cofactor function. These bacterial proteins are cofactor molecules that contribute to exosite formation and enhance the substrate presentation to the enzyme. At the same time, they modulate the specificity of plasmin towards substrates and inhibitors, making a 'specificity switch' possible.

Amino Acid Sequence↗
PubMed2000

Crystal structure of an in vivo HIV-1 protease mutant in complex with saquinavir: insights into the mechanisms of drug resistance.

Saquinavir is a widely used HIV-1 protease inhibitor drug for AIDS therapy. Its effectiveness, however, has been hindered by the emergence of resistant mutations, a common problem for inhibitor drugs that target HIV-1 viral enzymes. Three HIV-1 protease mutant species, G48V, L90M, and G48V/L90M double mutant, are associated in vivo with saquinavir resistance by the enzyme (Jacobsen et al., 1996). Kinetic studies on these mutants demonstrate a 13.5-, 3-, and 419-fold increase in Ki values, respectively, compared to the wild-type enzyme (Ermolieff J, Lin X, Tang J, 1997, Biochemistry 36:12364-12370). To gain an understanding of how these mutations modulate inhibitor binding, we have solved the HIV-1 protease crystal structure of the G48V/L90M double mutant in complex with saquinavir at 2.6 A resolution. This mutant complex is compared with that of the wild-type enzyme bound to the same inhibitor (Krohn A, Redshaw S, Richie JC, Graves BJ, Hatada MH, 1991, J Med Chem 34:3340-3342). Our analysis shows that to accommodate a valine side chain at position 48, the inhibitor moves away from the protease, resulting in the formation of larger gaps between the inhibitor P3 subsite and the flap region of the enzyme. Other subsites also demonstrate reduced inhibitor interaction due to an overall change of inhibitor conformation. The new methionine side chain at position 90 has van der Waals interactions with main-chain atoms of the active site residues resulting in a decrease in the volume and the structural flexibility of S1/S1' substrate binding pockets. Indirect interactions between the mutant methionine side chain and the substrate scissile bond or the isostere part of the inhibitor may differ from those of the wild-type enzyme and therefore may facilitate catalysis by the resistant mutant.

Amino Acid Substitution↗
PubMed2000

Crystal structure of the catalytic domain of human bile salt activated lipase.

Bile-salt activated lipase (BAL) is a pancreatic enzyme that digests a variety of lipids in the small intestine. A distinct property of BAL is its dependency on bile salts in hydrolyzing substrates of long acyl chains or bulky alcoholic motifs. A crystal structure of the catalytic domain of human BAL (residues 1-538) with two surface mutations (N186D and A298D), which were introduced in attempting to facilitate crystallization, has been determined at 2.3 A resolution. The crystal form belongs to space group P2(1)2(1)2(1) with one monomer per asymmetric unit, and the protein shows an alpha/beta hydrolase fold. In the absence of bound bile salt molecules, the protein possesses a preformed catalytic triad and a functional oxyanion hole. Several surface loops around the active site are mobile, including two loops potentially involved in substrate binding (residues 115-125 and 270-285).

Amino Acid Sequence↗
PubMed1999

Crystal structure of streptokinase beta-domain.

Streptokinase, a 47 kDa secreted protein of hemolytic strains of streptococci, is a human plasminogen activator and contains three structural domains linked by flexible loops. We describe here the crystal structure of the isolated streptokinase middle (SKbeta) domain determined at 2.4 A resolution. Among the functionally important structural features is a putative binding site for a kringle domain of plasminogen located at the tip of a fully exposed hairpin loop. The distribution of genetically conserved residues of SKbeta is strongly correlated with their functions. The extensive interface of the SKbeta dimer suggests that such dimers may also exist in solution for free SKbeta.

Amino Acid Sequence↗
PubMed1999

[Genetics and expression stability of exogenous gene construct in transgenic mice].

Transgenic mice were produced by introduced exogene construct lambda 106, an expression construct of HBsAg gene directed by bovine alpha-S1 casein gene, with microinjection. Gene integration test with PCR-Southern hybridization, shows that the construct integration rate is 56% (17/30) and expressed rate, by ELISA, of target gene product HBsAg is 100% (8/8) in the first generation. Generation trailing test suggests that the gene construct can be stably inherited across generation after generation, and expressed more or less than parents among offspring. However positive rate of offspring does not follow genetic ratio that exogenous gene can be integrated randomly into a single site on chromosomes. There may be a machinery of multi-sites for exo-gene integration. Changes of the target gene expression in the offsprings may be related to position effect, changing of gene copy number and changing of chromatin imprinting pattern when parent's genome information was delivered to their progeny.

Animals↗
PubMed1998

Crystal structure of the catalytic domain of human plasmin complexed with streptokinase.

Streptokinase is a plasminogen activator widely used in treating blood-clotting disorders. Complexes of streptokinase with human plasminogen can hydrolytically activate other plasminogen molecules to plasmin, which then dissolves blood clots. A similar binding activation mechanism also occurs in some key steps of blood coagulation. The crystal structure of streptokinase complexed with the catalytic unit of human plasmin was solved at 2.9 angstroms. The amino-terminal domain of streptokinase in the complex is hypothesized to enhance the substrate recognition. The carboxyl-terminal domain of streptokinase, which binds near the activation loop of plasminogen, is likely responsible for the contact activation of plasminogen in the complex.

Binding Sites↗
PubMed1997

The crystal structure of bovine bile salt activated lipase: insights into the bile salt activation mechanism.

BACKGROUND: The intestinally located pancreatic enzyme, bile salt activated lipase (BAL), possesses unique activities for digesting different kinds of lipids. It also differs from other lipases in a requirement of bile salts for activity. A structure-based explanation for these unique properties has not been reached so far due to the absence of a three-dimensional structure. RESULTS: The crystal structures of bovine BAL and its complex with taurocholate have been determined at 2.8 A resolution. The overall structure of BAL belongs to the alpha/beta hydrolase fold family. Two bile salt binding sites were found in each BAL molecule within the BAL-taurocholate complex structure. One of these sites is located close to a hairpin loop near the active site. Upon the binding of taurocholate, this loop becomes less mobile and assumes a different conformation. The other bile salt binding site is located remote from the active site. In both structures, BAL forms similar dimers with the active sites facing each other. CONCLUSIONS: Bile salts activate BAL by binding to a relatively short ten-residue loop near the active site, and stabilize the loop in an open conformation. Presumably, this conformational change leads to the formation of the substrate-binding site, as suggested from kinetic data. The BAL dimer observed in the crystal structure may also play a functional role under physiological conditions.

Animals↗
PubMed1997

The evaluation of developmental toxicity of chemicals exposed occupationally using whole embryo culture.

The purpose of this study was to employ the whole embryo culture (WEC) system to evaluate the developmental toxicity of industrial chemicals. Five chemicals including lead, cadmium, vinyl chloride, 1,2-dichloroethan, and carbon disulphide were tested in our laboratory both in vitro and in vivo (except lead). In vitro studies showed that cadmium and lead were teratogenic in the rat; whilst carbon disulphide, 1,2-dichloroethan and vinyl chloride mainly induced embryo growth retardation. The in vitro effects on development of the five industrial chemicals were similar to the effects in vivo. The in vitro effects were studied by three different exposure routes, direct exposure--chemicals added to the culture medium; indirect exposure--serum prepared from treated rats then used as culture medium, and pre-exposure--embryos treated maternally then explanted into control (untreated) culture medium. Comparing these three different exposure routes suggests that the last exposure route is the most effective when using WEC to evaluate developmental toxicity of industrial chemicals. The effects on embryo development of culturing in sera prepared from subjects occupationally exposed to antineoplastic drugs (ADs) was also tested by the WEC system. Embryos were cultured with human serum that was thought to contain ADs or ADs' metabolic materials (serum taken from nurses routinely handling ADs), to evaluate the effects of ADs on embryo development. Embryos (9.5-day) cultured with serum from 11 female nurses who had been handling ADs for 2-17 years in the oncology department all survived, but showed slight growth retardation. Embryos cultured with serum from 30 healthy and unexposed people served as controls and embryo development in their serum was normal.

Animals↗
PubMed1996

A cohort study of cancer among benzene-exposed workers in China: overall results.

A large cohort study of 74,828 benzene-exposed and 35,805 unexposed workers employed between 1972 and 1987 in 12 cities in China were followed to determine mortality from all causes and the incidence of lymphohematopoietic malignancies and other hematologic disorders. Benzene-exposed study subjects were employed in a variety of occupations, including painting, printing, and the manufacture of footwear, paint, and other chemicals. All-cause mortality was similar in the benzene-exposed and unexposed comparison group. Statistically significant excess deaths were noted among benzene-exposed subjects for leukemia (RR = 2.3, 95% CP 1.1-5.0), malignant lymphoma (RR = 4.5, 95% CI: 1.3-28.4), and nonneoplastic diseases of the blood (RR = 95% CP 2.5-infinity), and a marginally significant excess was noted for lung cancer (RR = 1.4, 95% CI: 1.0-2.0). Risk was significantly elevated for the incidence of all lymphohematopoietic malignancies (RR = 2.6, 95% CI: 1.5-5.0), malignant lymphoma (RR = 3.5, 95% CI: 1.2-14.9), and leukemia (RR = 2.6, 95% CI.. 1.3-5.7). Among the leukemia subtypes, only acute myelogenous leukemia (AML) incidence was significantly elevated (RR = 3.1, 95% CI: 1.2-10.7), although nonsignificant excesses were also noted for chronic myelogenous leukemia (CML) (RR = 2.6, 95% CI: 0.7-16.9) and lymphocytic leukemias (RR = 2.8, 95% CI.. 0.5-54.5). Significant excesses were found for aplastic anemia (RR = infinity, 95% CI: 2.2-co) and myelodysplastic syndrome (RR = infinity, 95% CI: 1.7-infinity). Employment in benzene-associated occupations in China is associated with a wide spectrum of myelogenous and lymphocytic malignant diseases and related disorders. Investigations continue to assess the nature of these associations.

Adult↗
PubMed1996

Indirect validation of benzene exposure assessment by association with benzene poisoning.

We present a validation study of a quantitative retrospective exposure assessment method used in a follow-up study of workers exposed to benzene. Assessment of exposure to benzene was carried out in 672 factories in 12 cities in China. Historical exposure data were collected for 3179 unique job titles. The basic unit for exposure assessment was a factory/work unit/job title combination over seven periods between 1949 and 1987. A total of 18,435 exposure estimates was developed, using all available historical information, including 8477 monitoring data. Overall, 38% of the estimates were based on benzene monitoring data. The highest time-weighted average exposures were observed for the rubber industry (30.7 ppm) and for rubber glue applicators (52.6 ppm). Because of its recognized link with benzene exposure, the association between a clinical diagnosis of benzene poisoning and benzene exposure was evaluated to validate the assessment method that we used in the cohort study. Our confidence in the assessment method is supported by the observation of a strong positive trend between benzene poisoning and various measures, especially recent intensity of exposure to benzene.

Benzene↗
PubMed1995

Equal vibrotactile sense thresholds of the fingers and its diagnostic significance for hand-arm vibration syndrome.

The vibrotactile sense thresholds (VSTs) of the middle fingers of 60 healthy persons and 97 patients with Hand-Arm Vibration Syndrome (HAVS) or subclinical HAVS were measured quantitatively. Intermittent vibratory irritations were adopted, with vibration stimulus frequencies at 8, 16, 31.5, 63, 125, 250, and 500 Hz. The equal VST contours of the fingers were mapped. Results showed that the VSTs of the normal group were not correlated with sex or handedness. From 8 Hz to 250 Hz the equal VST contours of the normal group were relatively flat; at more than 250 Hz the contours began an abrupt ascent. The VST values had a logarithmic rising tendency with the increasing age of subjects. In the equal VST contours the frequency of the most sensitive threshold value was 125 Hz in the normal group and 8 Hz in the HAVS group. The patients' VST values were higher than that of the healthy persons. The vibrotactilegram showed that the VST values of the patient groups first shifted at high frequencies and VST loss displayed a "V"-type hollow at 125 Hz and 250 Hz. The quantitative test method of VST was a valuable auxiliary detection method for HAVS. The "V"-type hollow of VST was an early clinical manifestation of HAVS.

Adult↗
PubMed1995

Characterization of a dinucleotide repeat in the 92 kDa type IV collagenase gene (CLG4B), localization of CLG4B to chromosome 20 and the role of CLG4B in aortic aneurysmal disease.

Proteolytic imbalance may play a role in the pathogenesis of abdominal aortic aneurysms (AAA). CLG4B, which encodes the 92-kDa form of type IV collagenase, is a candidate gene for AAA. We genotyped a polymorphic dinucleotide repeat in the 5' flanking region of CLG4B in 94 unrelated Caucasian controls and in 127 unrelated Caucasian AAA cases. Eight alleles were detected in 188 control chromosomes with an observed heterozygosity of 0.68. There was no significant difference in allele distribution between cases and controls. We genotyped the dinucleotide repeat in 10 CEPH reference pedigrees and performed pairwise linkage analysis with markers on each of the 22 human autosomes. Lod scores between 10.45 and 20.29 were observed with markers spanning chromosome region 20q11.2-q13.1. Further support for assignment of CLG4B to chromosome 20 was provided by analysis of human-rodent somatic cell hybrids. This work describes a highly polymorphic marker in the CLG4B gene and assigns this gene to chromosome 20.

Aortic Aneurysm↗
PubMed1994

[Determination of finger vibrotactile threshold and its significance in patients with hand-arm vibration syndrome].

Vibrotactile sense threshold (VST) of the end of the middle fingers in 97 patients with established or suspected hand-arm vibration syndrome and 60 healthy controls was determined, and finger VST contour and vibration damage curve were plotted. Results revealed the most sensitive frequencies of VST were 8 Hz for the patients and 125 Hz for the controls, respectively. VST in the patients with hand-arm vibration syndrome was higher than that in the controls, and the threshold shift was initially at high frequency with a "V" dip at 125 Hz and 250 Hz. So, the authors think determination of VST is an auxiliary method used for diagnosis of hand-arm vibration syndrome.

Adult↗
PubMed1993

[The physical properties, pharmacology and antifertility action of crystal trichosanthin protein liposome].

The crystal trichosanthin protein liposome (TPL) was prepared by an emulsion-forming method. The rate of entrapment was 33.0-44.8% and the rate of recovery was 95.15-99.58%. The determination of trichosanthin protein was accomplished by spectrophotometry at 650 nm, thus eliminating completely the interference of lipids. The distribution of particle size of TPL was mainly in the range of 1-2 microns (96.9%). TPL was sterilized by 60Co radiation at 4 degrees C for 90 min and was stable when stored at 4 degrees C, 25 degrees C and 37 degrees C. The seepage rate of trichosanthin from the liposome was 15.22% after storage for 38 days at room temperature. Experiments showed that TPL had a slight allergic reaction, but was safe by hemolysis and coagulation tests. When the mice were given TPL ip 8 micrograms per animal 6 days after pregnancy, the rate of termination of early pregnancy was 83.3%. It thus appears that TPL can terminate early pregnancy in mice distinctly. This indicates that TPL may be expected to be developed into a new type of anti-early pregnancy preparation.

Abortion, Induced↗