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Biomedical subjects

Y Xu

Publications and source records attributed to Y Xu.

At least 1,045 records · Page 58Linked to original sources
PubMed1994

Epidemiological and genetic studies of congenital profound deafness in the general population of Sichuan, China.

People with congenital profound deafness (CPD) were surveyed in the general population of Sichuan. The prevalence was 0.082% (104/126, 876) of the general population, male 0.086%, female 0.078%. There was no significant difference in prevalence between urban and rural populations. However the population living in the mountains had a much higher prevalence than people from the foothills and plains (P < 0.05). Amongst the nationalities investigated, there were significant differences in prevalence. Non-Chinese, except for Tibetans, presented a significantly higher prevalence than Han Chinese. There was a significantly higher prevalence among the inbred population (0.82%) than among the non-inbred population (0.72%). An effect of parental age was demonstrated, but no effect of birth order was found. There was a significant seasonal variation in prevalence. Inherited cases could account for 71.2% of all cases, of which 92% were autosomal recessive (AR) and 8% autosomal dominant (AD). Heterogeneity in AR was found with at least 8 different loci. The fitness was 60.26%, the coefficient of selection was 0.3974, mutation rate was estimated to be 2.0 x 10(-4), and no heterozygote advantage was proven.

Adolescent↗
PubMed1994

Enhancement of performance in sedimentation field-flow fractionation by temperature elevation.

Nonequilibrium theory, combined with the principles of time optimization, show that the time necessary to achieve a given separation in FFF is scaled to eta/T, where eta is viscosity and T is absolute temperature. The eta/T ratio for water (and other common liquids) decreases several-fold for modest temperature gains of approximately 20-60 degrees C, implying a significant advantage for FFF operation at elevated temperatures. This concept was tested by modifying a standard sedimentation FFF apparatus with a heating system. The separation of 0.220-0.742 microns polystyrene latex beads in aqueous carrier liquids was compared at room temperature and at elevated temperatures of 51 and 68 degrees C. Both separation power and speed were improved. In accordance with the predicted eta/T scaling, the separation time of five bead sizes at a given resolution level was reduced by a factor of approximately 2.4 (from 29 to 12 min) in elevating the temperature from 25 to 68 degrees C. Some other potential benefits of temperature elevation in FFF are discussed.

Chemical Fractionation↗
PubMed1994

Relative specificities of a series of beta-lactam-recognizing enzymes towards the side-chains of penicillins and of acyclic thioldepsipeptides.

In an attempt to understand more of the subtle differences between bacterial beta-lactamases and DD-peptidases, comparisons have been made between the specificities of these enzymes towards the phenylacetyl side chain, generally thought to be favoured by beta-lactamases, and the NN'-diacetyl-L-lysyl side chain, widely employed in low-molecular-mass substrates of DD-peptidases. These comparisons were carried out with both a penicillin and an acyclic thioldepsipeptide reaction nucleus and employing a range of both beta-lactamases and DD-peptidases. Rather contrary to general expectations, a general preference for reaction of both groups of enzymes with penicillins rather than thioldepsipeptides was observed and for the phenylacetyl rather than the NN'-diacetyl-L-lysyl side chain. Quantitative comparisons suggested that the side chains of penicillins may be bound in relatively similar sites in all of the enzymes whereas the side chains of thioldepsipeptides are more heterogeneously bound, both with respect to each other and to the comparable side chains of penicillins.

Binding Sites↗
PubMed1994

Somatostatin regulates somatostatin receptor subtype mRNA expression in GH3 cells.

Homologous up-regulation of somatostatin receptors (SSTR) was investigated in the GH3 clonal pituitary cell line. We report that chronic exposure to SRIF which has high affinity for all SSTR subtypes leads to a net increase in receptor binding and SSTR subtype mRNA. Treatment of cells with 1 microM SRIF increased specific [125I-Tyr11]SRIF binding to 280% of control values by 24 hr and to 350% by 48hr. Associated with the increase in SSTR binding was a net increase in SSTR subtype mRNA expression. Levels of SSTR1 increased to over 400% of control values by 6 hr and remained elevated for up to 48 hr. SSTR3, SSTR4, and SSTR5 mRNA was also dramatically increased at 24 and 48 hr. SSTR2 mRNA, in contrast, showed a biphasic response, initially increasing to 150% of control at 2 hr then decreasing to 50% at 6 hr with normalization by 48 hr. Our data suggests that multiple mechanisms contribute to the highly regulated expression of SSTR subtypes. The identification of specific molecular and cellular mechanisms mediating homologous SSTR up-regulation in GH3 cells and the involvement this process has in mediating the diverse biological effects of somatostatin are topics of current research.

Animals↗
PubMed1994

The crystal structure of human hypoxanthine-guanine phosphoribosyltransferase with bound GMP.

The crystal structure of HGPRTase with bound GMP has been determined and refined to 2.5 A resolution. The enzyme has a core alpha/beta structure resembling the nucleotide-binding fold of dehydrogenases, and a second lobe composed of residues from the amino and carboxy termini. The GMP molecule binds in an anti conformation in a solvent-exposed cleft of the enzyme. Lys-165, which forms a hydrogen bond to O6 of GMP, appears to be critical for determining the specificity for guanine and hypoxanthine over adenine. The location of active site residues also provides evidence for a possible mechanism for general base-assisted HGPRTase catalysis. A rationalization of the effects on stability and activity of naturally occurring single amino acid mutations of HGPRTase is presented, including a discussion of several mutations at the active site that lead to Lesch-Nyhan syndrome.

Amino Acid Sequence↗
PubMed1994

Kinetics of biotinyl-5'-adenylate synthesis catalyzed by the Escherichia coli repressor of biotin biosynthesis and the stability of the enzyme-product complex.

The Escherichia coli repressor of biotin biosynthesis is both a biotin ligase and the repressor of transcriptional initiation at the biotin biosynthetic operon. The small molecule, biotinyl-5'-adenylate (bio-5'-AMP), is the intermediate in the biotin ligation reaction and the positive allosteric effector for sequence-specific DNA binding by BirA. Synthesis of the adenylate from the substrates biotin and ATP is catalyzed by BirA. Although BirA and other biotin holoenzyme synthetases have been the subject of biochemical studies, no direct measurements of the bio-5'-AMP synthesis reaction have been reported. No information relating to the mechanism and kinetic parameters governing adenylate synthesis is available. In addition to this lack of kinetic information, the thermodynamic stability of the BirA-bio-5'-AMP complex is not known. Since the BirA-adenylate complex plays a pivotal role in the biotin regulatory system, both the kinetic and thermodynamic information are essential to a quantitative understanding of the system. We have developed a method for measuring the time course of bio-5'-AMP synthesis. The results of these measurements indicate that the time course is characterized by an initial burst followed by a slow linear phase. The burst corresponds to the rapid synthesis of 1 mol of product per mole of enzyme, and the rate of the slow linear phase is limited by the release of product from the enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Monophosphate↗
PubMed1994

19F NMR calcium changes, edema and histology in neonatal rat brain slices during glutamate toxicity.

Respiring neonatal cerebrocortical slices (350 microns thick), loaded with the free calcium indicator 5F-BAPTA, were perfused in a 20-mm-diameter glass NMR tube with oxygenated artificial CSF, exposed to extracellular glutamate and studied at 4.7 Tesla with 19F NMR spectroscopy. 31P/1H NMR spectra, obtained concurrently, were used to assess slice integrity from determinations of intracellular pH, ATP, PCr, lactate and N-acetylaspartate. 60-min periods were induced of recoverable and nonrecoverable glutamate toxicity-defined from changes in NMR metabolites. In other NMR studies, where 5F-BAPTA was not used, metabolic toxicity was modulated by three glutamate receptor antagonists: dizocilpine, NBQX and kynurenic acid. Outcome measurements were made of edema, determined invasively in isolated slices from % swelling and water content and from histological changes in Nissl stains of slice sections. Edema was (1) detectable in all slices within minutes after onset of glutamate exposure, though never in untreated control slices, and (2) modulated differently by dizocilpine, NBQX and kynurenate. Correlations were observed between edema and NMR decreases in PCr and ATP. Nissl stains of sections from slices treated with the most protective agent, dizocilpine, showed preservation of neuronal processes. As was expected in 7-day-old rats with immature NMDA receptors, 19F NMR spectroscopy revealed only small increases in free intracellular calcium ([Ca2+]i). These occurred late during glutamate exposure and reversed early during glutamate washout. The studies demonstrate that it is possible to study correlations between repeated noninvasive NMR spectra in ensembles of brain slices and invasive measures of early cellular responses.

Animals↗
PubMed1994

Mutational analysis of the PRP4 protein of Saccharomyces cerevisiae suggests domain structure and snRNP interactions.

The PRP4 protein of Saccharomyces cerevisiae is an essential part of the U4/U6 snRNP, a component of the mRNA splicing apparatus. As an approach to the determination of structure-function relationships in the PRP4 protein, we have isolated more than fifty new alleles of the PRP4 gene through random and site-directed mutagenesis, and have analyzed the phenotypes of many of them. Twelve of the fourteen single-point mutations that give rise to temperature-sensitive (ts) or null phenotypes are located in the portion of the PRP4 gene that corresponds to the beta-transducin-like region of the protein; the remaining two are located in the central portion of the gene, one of them in an arginine-lysine-rich region. Nine additional deletion or deletion/insertion mutations were isolated at both the amino- and carboxy-termini. These data show that the amino-terminal region (108 amino acids) of PRP4 is non-essential, while the carboxy-terminal region is essential up to the penultimate amino acid. A deletion of one entire beta-transducin-like repeat (the third of five) resulted in a null phenotype. All ts mutants show a first-step defect in the splicing of U3 snRNA primary transcript in vivo at the non-permissive temperature. The effects on prp4 mutant growth of increased copy-number of mutant prp4 genes themselves, and of genes for other components of the U4/U6 snRNP (PRP3 and U6 snRNA) have also been studied. We suggest that the PRP4 protein has at least three domains: a non-essential amino-terminal segment of at least 108 amino acids, a central basic region of about 140 residues that is relatively refractile to mutation and might be involved in RNA interaction, and an essential carboxy-terminal region of about 210 residues with the five repeat-regions that are similar to beta-transducins, which might be involved in protein-protein interaction. A model of interactions of snRNP components suggested by these results is presented.

Amino Acid Sequence↗
PubMed1994

Distribution and chromatographic analysis of galanin message-associated peptide (GMAP)-like immunoreactivity in the rat.

Galanin message-associated peptide (GMAP) constitutes the C-terminal part of the precursor protein encoding also the biologically active neuropeptide galanin (GAL). We have raised antisera against a species-conserved portion of GMAP, and investigated the localization of GMAP-like immunoreactivity (-LI) in relation to that of GAL-LI in the rat central and peripheral nervous system using the indirect immunofluorescence technique. In the central nervous system, GMAP-immunoreactive (-IR) cell bodies were observed in the hypothalamus, while GMAP-IR nerve fibers were demonstrated in the septum, hypothalamus, pons and spinal cord. In the posterior pituitary and in the connecting infundibular stalk, weakly fluorescent GMAP-IR nerve fibers were observed. GMAP-IR nerve fibers were also observed throughout the gastrointestinal tract, i.e., from the stomach down to the colon, and in all layers, except in the epithelium, of the wall. In general, the staining of consecutive tissue sections suggested that GMAP-IR was co-distributed with that of GAL-IR. A sensitive radioimmunoassay (RIA) for characterization of GMAP-IR in the rat central and peripheral nervous system was also developed. Characterization of GMAP-LI in acid extracts of rat brain and small intestine, using reverse phase high pressure liquid chromatography (rpHPLC), revealed multiple GMAP-IR forms that co-eluted with a synthetic porcine GMAP(19-41)-amide fragment, or were less or more polar than this fragment. The corresponding chromatographic analysis of GAL-LI revealed only one major form corresponding to rat GAL. The immunohistochemical data indicate that a GMAP-like peptide(s) probably is axonally transported and may possibly have pre- and/or post-synaptic functions. The nature of the multiple GMAP-IR components remains to be investigated, but may tentatively represent differently processed and/or chemically modified forms of rat GMAP(1-60).

Amino Acid Sequence↗
PubMed1994

Role of histidine 35 of the S1 subunit of pertussis toxin in the ADP-ribosylation of transducin.

Molecular modeling of the S1 subunit (S1) of pertussis toxin with other ADP-ribosylating bacterial exotoxins predicted that histidine 35 (His-35) would residue within the active site of S1. Recombinant derivatives of S1 (rS1 and the C180 peptide) which contained either a H35Q or H35P mutation were analyzed to determine the role of His-35 in ADP-ribosylation. C180 peptide is a recombinant peptide composed of the amino-terminal 180 amino acids of S1. Under linear velocity conditions, C180H35Q possessed 2% of wild type C180 peptide activity and C180H35P possessed no detectable activity in the ADP-ribosylation of transducin. The H35Q mutation did not change the affinity of recombinant peptides for NAD or two targets for ADP-ribosylation, transducin, or alpha i3C20, but did lower the kcat in the NAD glycohydrolase and ADP-ribosyltransferase reactions. Neither the H35Q nor H35P mutation reduced the ability of recombinant proteins to be photocross-linked with NAD which was consistent with the His-35 mutations not reducing the affinity for NAD. These data indicate that His-35 does not reduce the affinity of S1 for NAD or transducin, but functions as a catalytic residue in the ADP-ribosylation reaction possibly in a hydrogen bonding capacity.

Adenosine Diphosphate Ribose↗
PubMed1994

Biochemical analysis of mutations at tyrosine-98 of the S1 subunit of pertussis toxin.

Molecular modeling and alignment of the primary amino acid sequence of the S1 subunit (S1) of pertussis toxin (PT) with other members of the family of ADP-ribosylating bacterial exotoxins predicted that tyrosine-98 (Y98) of S1 was a conserved residue among these exotoxins. To extend our understanding of the relationship between pertussis toxin and the other ADP-ribosylating exotoxins, we defined the function of Y98 of S1. Using site-directed mutagenesis, Y98 of S1 was substituted with alanine (Y98A), leucine (Y98L), histidine (Y98H), and phenylalanine (Y98F). Mutations were analyzed in the C180 peptide and C219 peptide, recombinant derivatives of S1 which contain the first 180 and 219 amino-terminal residues of S1, respectively. Periplasmic extracts containing the Y98n peptides expressed similar specific activities for the ADP-ribosylation of transducin (Gt) as the periplasmic extract containing wild-type peptides. Mutations at Y98 influenced the subcellular localization of the respective Y98n peptide. The majority of the wild-type Y98 and Y98F peptides localized to the periplasmic extract, while the majority of Y98A and Y98L peptides were associated with the insoluble bacterial outer membrane. Purified C180Y98A and C180Y98F and partially purified C180Y98H peptides possessed similar specific activities for the ADP-ribosylation of Gt as the wild-type C180 peptide. KmNAD and kcat for C180Y98A and C180Y98F in the NAD glycohydrolase reaction were similar to the wild-type C180 peptide. These data show that the R group of Y98 does not participate in the ADP-ribosylation of Gt, but appears to contribute to the proper folding of S1.

Adenosine Diphosphate Ribose↗
PubMed1994

Energetics of pyruvate phosphate dikinase catalysis.

The present study was carried out to determine the energetics of Clostridium symbiosum pyruvate phosphate dikinase (PPDK) catalyzed interconversion of adenosine 5'-triphosphate (ATP), orthophosphate (Pi), and pyruvate (pyr) with adenosine 5'-monophosphate (AMP), inorganic pyrophosphate, and phosphoenolpyruvate (PEP) [E.ATP <==> E-PP.AMP <==> E-PP.AMP.Pi <==> E-P.AMP.PPi <==> E-P.pyr <==> E.PEP where E-PP and E-P represent the pyrophosphoryl and phosphoryl enzyme intermediates]. Thermodynamic techniques were used along with steady-state and pre-steady-state kinetic techniques to determine the rate constants for the substrate/product binding and release steps and the rate constants for the forward and reverse chemical steps. These values were used along with estimates of the cellular concentrations of the substrates and products to construct the free energy profile for the enzymatic reaction under physiological conditions. The energy profile obtained with the Mg2+/NH4(+)-activated enzyme revealed well-balanced transition states and well-balanced internal ground state energies (i.e., within 1 kcal/mol of each other). Examination of the energetics of the reaction steps leading from ATP to phosphohistidine formation in E-P suggested the use of intrinsic binding energy in the synthesis of a high energy P-N linkage. Comparison of the energy profiles of the Mg2+/NH4(+)-vs Co2+/NH4(+)-activated enzymes revealed cofactor selectivity at each of the phosphosphoryl transfer steps.

Adenosine Monophosphate↗