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Q J Zhang

Publications and source records attributed to Q J Zhang.

15 recordsLinked to original sources
PubMed1992

Immune escape by Epstein-Barr virus (EBV) carrying Burkitt's lymphoma: in vitro reconstitution of sensitivity to EBV-specific cytotoxic T cells.

Epstein-Barr virus (EBV) positive Burkitt's lymphoma (BL) cells are markedly less sensitive to EBV-specific cytotoxic T cell (CTL) recognition than EBV-transformed lymphoblastoid cell lines of normal B cell origin. Three features of the BL cell phenotype might contribute to this reduced susceptibility: (i) low expression of cell adhesion molecules, (ii) low expression of HLA class I and selective down-regulation of particular alleles, and (iii) down-regulation of all transformation-associated EBV antigens except EBV-encoded nuclear antigen (EBNA)-1. This study assesses the individual importance of each of these features for immune escape. For this purpose the WW1-BL cell line was used which expresses all the known transformation-associated EBV antigens (EBNA-1 to -6 and latent membrane protein-1 and -2) but which is negative for HLA A11 and for the adhesion molecule leukocyte function associated antigen-3 (LFA-3). Using recombinant vectors, these deficiencies have been sequentially corrected and the cells have been tested for sensitivity to EBV (B95.8 strain)-induced CTL preparations recognizing epitope(s) of EBNA-4 in the context of HLA A11. Expression of HLA A11 alone or in combination with LFA-3 did not sensitize WW1-BL cells to these effectors. Lysis was only achieved when HLA A11 was co-expressed with the B95.8 virus-encoded EBNA-4 protein, and in these circumstances sensitization did not require LFA-3. These results indicate that reconstitution of the relevant HLA-EBV epitope target complex on the cell membrane is sufficient to render BL cells sensitive to virus-specific cytolysis. The requirement for EBNA-4 reconstitution to achieve lysis of the WW1-BL/A11 transfectant suggested that the resident WW1 virus-encoded EBNA-4 protein did not contain the relevant target epitope for HLA A11-restricted recognition. This was confirmed by transferring the WW1 virus isolate into another A11-positive B cell background.

Antigens, CD↗
PubMed1992

[Studies on the function of Ser579 and Arg580 in beta-subunit of penicillin G acylase with the method of site-specific mutagenesis].

According to the comparison of amino acid sequence between PGA (Penicillin G Acylase) and PBPs (Penicillin Binding Protein), We suggest that No. 565-595 peptide fragment in beta-subunit of PGA may be a substrate-binding site of enzyme. Plasmid pTZGA was constructed by cloning the 2.6 kb PGA gene of pWGA into phagemid pTZ18U The technique of site-specific mutagenesis was used to study the role of residue No. 579 (Ser) and No. 580 (Arg) of PGA. Four kinds of mutants were obtained (Ser579-->Gly579, Arg580-->Gly580, Arg580-->Glu580, Arg580-->Lys580), both Glu580 and Gly580 mutants showed no activity of enzyme and Lys580 mutant remained 30% and Gly579 mutant kept 70% activity of wilde type. The same protein expression of four mutants according to the results of ELISA indicate that mutation does not affect the expression of PGA, but Arg580 residue may be essential for substrate-binding or catalysis of PGA.

Amino Acid Sequence↗
PubMed1991

2-Nitro-5-(6-bromohexanoylamino)benzoic acid test paper method for detecting microorganisms capable of producing cephalosporin acylases.

A novel method for detecting microorganisms capable of producing cephalosporin C (CPC) acylase and/or 7-(4-carboxybutanamido)cephalosporanic acid (GL-7-ACA) acylase has been developed. The method is based on the degradation of 2-nitro-5-(6-bromohexanoylamino)benzoic acid (NBHAB), a chromogenic substrate, into yellow 2-nitro-5-aminobenzoic acid by the action of the CPC acylase or the GL-7-ACA acylase. This method is very sensitive and quite specific, and has been successfully applied to screen the acylases from a variety of bacteria. A large number of colonies isolated on a plate surface from more than 67 samples and several known bacteria were tested by the NBHAB paper. Five NBHAB-positive strains and isolates were obtained. They were further examined by the reaction of their bacterial cells upon CPC and GL-7-ACA, respectively, and by thin-layer chromatography in order to distinguish the CPC acylase from the GL-7-ACA acylase.

Agar↗
PubMed1991

Cloning and expression of penicillin G acylase gene in Bacillus megaterium.

Bacillus megaterium BM1, which produces penicillin G acylase (PGA), has been isolated. Gene encoding for PGA was cloned into E. coli MC1061 using pBR322 as the vector, obtaining a recombinant plasmid pBmPA4 containing 9.9 kb inserted DNA. Restriction map of the plasmid was analyzed. A pBmPA5 containing 4.9 kb was gained by deletion in vitro. Both pBmPA4 and pBmPA5 clones can be expressed in E.coli MC1061, and their expressions were induced by phenylacetic acid.

Bacillus megaterium↗
PubMed1991

Threshold of transmission of Brugia malayi by Anopheles sinensis.

In an area of central China where Brugia malayi is transmitted by Anopheles sinensis, three villages were followed for 4 years without any control measures. Diethylcarbamazine (DEC) had been used in a control programme reducing the parasite rate in Shuiwa from 12.72 to 0.59%, in Gubomen from 3.18 to 1.55% and in Moshi from 5.62 to 2.81% (although this fell the following year to 2.23%) up until the start of the trial. The village populations, the human biting rate, the parous and the gonotrophic cycle were comparable for all the areas, yet the microfilariae rate dropped to 0.18% in Shuiwa, 0.31% in Gubomen, but not in Moshi where it increased from 2.23 to 2.43%. This indicated that Shuiwa and Gubomen were below the threshold of transmission. Due to the small number of positive cases remaining, the density measurements showed less change, but a separate study on 44 individuals followed for 6 years revealed that those above 13 microfilariae per 60 mm3 remained unchanged or increased, while those below progressively decreased their densities. The threshold of transmission of B. malayi by An. sinensis was considered to be between 1.55 and 2.23% of the population, providing no individual had a higher density than 12 microfilariae per 60 mm3.

Animals↗
PubMed1991

[Effect of mutagenesis at Ser 177 residue in penicillin G acylase on activity of the enzyme].

The technique of cassette and site-specific mutagenesis were used to study the role of residue No. 177 in penicillin G acylase (PGA, EC 3.5.1.11). Ser is conserved at residue No. 177 in all penicillin binding proteins. We got a series of mutants in which the amino acid at residue No. 177 was replaced by other amino acids through the site-specific and cassette mutagenesis, and we characterized the mutants by colony hybridization, NIPAB paper test and DNA sequence analysis. These mutants all show no activity of enzyme, even if the Ser residue was replaced by Thr, Gly and Ala respectively. The results show that Ser residue may be essential for substrate-binding or catalysis of PGA.

Escherichia coli↗
PubMed1990

A novel method for isolation and concentration of ribonuclease T1 from Taka-Diastase.

A novel method for isolation and concentration of RNase T1 from Taka-Diastase is developed. It is a combination method of bentonite adsorption with dialysis desorption. In the present method, RNase T1 can be concentrated about ten-fold, the recovery of total activity was greater than 95%, and specific activity was raised 8-10 folds. Further purification with ammonium sulfate precipitation and chromatography on DEAE-cellulose and DEAE-Sephadex yields a RNase T1 which contains no pMase. pDase nor RNase T2 activities and a 750 fold increase in specific activity. Our method is more simple, rapid, and efficient than previous methods.

Ammonium Sulfate↗
PubMed1990

Sequence analysis of D-xylose isomerase gene from Escherichia coli.

Gene coding for E. coli D-xylose isomerase was cloned into the plasmid pWR13 from the plasmid pXO100 containing D-xylose operon. Several subclones harboring DNA fragments of different size obtained by step- by-step shortening with Bal31 nuclease were directly sequenced. The complete nucleotide sequence of the 1.6kb DNA fragment carrying D-xylose isomerase gene was determined. The structural gene of 1320 nucleotides codes for a protein of 440 amino acids. The additional 209 nucleotides at 5' end and 82 nucleotides at 3' end include Shine-Dalgarno ribosome-binding site and the transcriptional termination codon, respectively.

Aldose-Ketose Isomerases↗
PubMed1990

[Cloning and expression of E. coli glucose isomerase gene in Streptomyces lividans].

E. coli glucose isomerase gene was cloned into Streptomyces lividans using shuttle plasmid vector pSE-3. First plasmid pXI203 was constructed in E. coli using plasmids pXI200 (containing 1.6 kb glucose isomerase gene) and pGEM-3 digested with EcoR1. Then, recombinant plasmid pSEX100 was also constructed in E. coli using plasmids pXI203 and pSE-3 digested with HindIII. When the pSEX100 was transformed into Streptomyces lividans protoplasts, recombinants were obtained on R5YE medium containing 50 micrograms/ml neomycin and 50 micrograms/ml thiostrepton. The results showed that the E. coli glucose isomerase gene cloned and expressed in Streptomyces lividans via the analysis of restriction enzyme digestion as well as the detection of the glucose isomerase activity.

Aldose-Ketose Isomerases↗
PubMed1989

[Whole nucleotide sequence of penicillin G acylase gene and its flanking region from E. coli].

Some of microorganisms have been known to possess penicillin G acylase activity. The E. coli derived penicillin G acylase (PGA) can catalyze the conversion of penicillin G into phenylacetic acid and 6-amino-penicillanic acid, the latter is used as the starting compound for the industrial formation of semi-synthetic penicillins. Apart from its industrial importance, the enzyme PGA displays a number of interesting properties. Catalytically active enzyme is localized in the periplasmic space of E. coli cells and composed of two dissimilar subunits. The two subunits are apparently produced from a precursor protein, via a processing pathway hitherto unique in its features for a prokaryotic enzyme. The studies on processing of the precursor and on the relationship between structure and function of the mature enzyme are important theoretically. Previously we cloned a 3.5 kb DNA fragment from a strain (E. coli AS 1.76), which displays PGA activity. In this paper, we report a nucleotide sequence of the 3.5 kb DNA fragment containing PGA gene. After insertion of the DNA fragment into EcoR I and Hind III sites in pWR 13, pPGA 20 had been obtained. We subcloned the Hind III and Bg1 II treated fragment of 1.6 kb in length from pPGA 20 into Hind III and BamH I sites of pWR 13 to get a pPGA 1.6, and Bg1 II and EcoR I treated fragment of 1.9 kb in length into BamH I and EcoR I sites of pWR 13 to get a pPGA 1.9. The linearized pPGA 1.9 which were digested with appropriate restriction enzymes were progressively shortened from both ends respectively by digestion with Bal 31 nuclease, followed by cleavage of shortened target DNA off vector DNA molecules with appropriate restriction enzymes. The series of the DNA fragments shortened from EcoR I end were then cloned into plasmid pWR 13 which had previously digested with Hind III and Sma I enzymes (Fig. 1). The DNA fragment cloned in pWR 13 were directly sequenced on the resulted plasmids by using primer I and primer II. Thus we have obtained the complete nucleotide sequence of the 3.5 kb DNA fragment. The 3.5 kb fragment contains an intact PGA gene which is 2.6 kb.(ABSTRACT TRUNCATED AT 400 WORDS)

Amidohydrolases↗
PubMed1986

A method for screening penicillin G acylase-producing bacteria by means of 2-nitro-5-phenylacetaminobenzoic acid test paper.

A simple, rapid assay for screening penicillin G acylase-producing bacteria is presented. The method is based on the formation of yellow 2-nitro-5-aminobenzoic acid by penicillin G acylase acting on 2-nitro-5-phenylacetaminobenzoic acid (NIPAB). NIPAB test paper is briefly applied to bacterial colonies on the agar surface, which are subsequently scored individually on the paper by color; bright yellow indicates the presence of penicillin G acylase, natural color its absence. The present method is suitable not only for screening penicillin G acylase-production by a variety of bacteria but also for detection from a large number of transformant colonies of clones containing a gene encoding for the enzyme.

Amidohydrolases↗