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Feng Gong

Publications and source records attributed to Feng Gong.

17 recordsLinked to original sources
PubMed2006

Rad4-Rad23 interaction with SWI/SNF links ATP-dependent chromatin remodeling with nucleotide excision repair.

Chromatin rearrangement occurs during nucleotide excision repair (NER). Here we show that Snf6 and Snf5, two subunits of the SWI/SNF chromatin-remodeling complex in Saccharomyces cerevisiae, copurify with the NER damage-recognition heterodimer Rad4-Rad23. This interaction between SWI/SNF and Rad4-Rad23 is stimulated by UV irradiation. We demonstrate that NER in the transcriptionally silent, nucleosome-loaded HML locus is reduced in yeast cells lacking functional SWI/SNF. In addition, using a restriction enzyme accessibility assay, we observed UV-induced nucleosome rearrangement at the silent HML locus. Notably, this rearrangement is markedly attenuated when SWI/SNF is inactivated. These results indicate that the SWI/SNF chromatin-remodeling complex is recruited to DNA lesions by damage-recognition proteins to increase DNA accessibility for NER in chromatin.

Adenosine Triphosphate↗
PubMed2006

Decreased immunorejection in unmatched blood transfusions by attachment of methoxypolyethylene glycol on human red blood cells and the effect on D antigen.

BACKGROUND: Pegylation of red blood cells (RBCs) has been the primary focus of research on the immunocamouflage of cell. The aim of this study was to demonstrate pegylation homogeneity, its shielding effect on D antigens, and its storage stability. In addition, methoxypolyethylene glycol (mPEG)-modified RBCs (mPEG-RBCs) were tested serologically against a panel of serum samples that was difficult to match to find a solution to the difficulty in matching. STUDY DESIGN AND METHODS: In this study, fluorescein-PEG and a confocal laser scanning microscope were used to monitor PEG attachment on RBC population and observe reaction homogeneity, the stability of mPEG combined with RBCs in vitro was evaluated by the RBC ghost agglutination test, the pegylation sites on membrane were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis with two dye methods, and the effect of pegylation on D antigen was detected by immunoblotting techniques. Compatibility tests were carried out between 66 cases of serum with difficulty in blood matching and mPEG-camouflaged RBCs by use of four blood matching methods including direct agglutination, indirect antiglobulin test (IAT), microtyping gel cards (MTS), and the manual polybrene technique (MPT). RESULTS: The results indicated the homogeneity of pegylation, the absence of RhD protein in mPEG-modified D+ RBCs by Western blotting, and attachment of PEG to RBCs after 30 days of storage, while RBCs still remained antigenically silent. All pegylation RBCs showed a negative reaction with ABO-matched patients' serum samples by direct agglutination, IAT, and MTS, which indicated that pegylation RBCs and patients' serum samples were compatible. MPT was not suitable for detecting blood matching of mPEG-RBCs, because modification changed the RBCs' biophysical properties. CONCLUSION: In conclusion, mPEG-RBCs have acceptable in vitro properties and provide a useful solution to problems with clinical blood matching, although such masking leaves much to be desired.

Blood Grouping and Crossmatching↗
PubMed2006

Overexpression of tnaC of Escherichia coli inhibits growth by depleting tRNA2Pro availability.

Transcription of the tryptophanase (tna) operon of Escherichia coli is regulated by catabolite repression and tryptophan-induced transcription antitermination. Induction results from ribosome stalling after translation of tnaC, the coding region for a 24-residue leader peptide. The last sense codon of tnaC, proline codon 24 (CCU), is translated by tRNA(2)(Pro). We analyzed the consequences of overexpression of tnaC from a multicopy plasmid and observed that under inducing conditions more than 60% of the tRNA(2)(Pro) in the cell was sequestered in ribosomes as TnaC-tRNA(2)(Pro). The half-life of this TnaC-tRNA(2)(Pro) was shown to be 10 to 15 min under these conditions. Plasmid-mediated overexpression of tnaC, under inducing conditions, reduced cell growth rate appreciably. Increasing the tRNA(2)(Pro) level relieved this growth inhibition, suggesting that depletion of this tRNA was primarily responsible for the growth rate reduction. Growth inhibition was not relieved by overexpression of tRNA(1)(Pro), a tRNA(Pro) that translates CCG, but not CCU. Replacing the Pro24CCU codon of tnaC by Pro24CCG, a Pro codon translated by tRNA(1)(Pro), also led to growth rate reduction, and this reduction was relieved by overexpression of tRNA(1)(Pro). These findings establish that the growth inhibition caused by tnaC overexpression during induction by tryptophan is primarily a consequence of tRNA(Pro) depletion, resulting from TnaC-tRNA(Pro) retention within stalled, translating ribosomes.

Escherichia coli↗
PubMed2006

[Preliminary study on xenotransfusion from porcine red blood cell into Rhesus monkey].

In order to study the possibility of xenotransfusion from porcine red blood cell (pRBC) to primate, the antigens on pRBC surface were modified to make it more compatible to primate sera. Porcine RBCs were subjected to both enzymatic removal of membrane alpha-Gal antigens with recombinant alpha-galactosidase (AGL) and covalent attachment of succinimid propionate-linked methoxypolyethyleneglycol (mPEG-SPA) to camouflage non-alphaGal antigens. The effects of double modifications were determinated by hemagglutination and clinical cross-match testing with rhesus sera. In vivo clearance rates and safety of modified pRBCs were measured after it was transfused into Rhesus monkey with or without immunosuppressant treatment. The validity of pRBC was detected in exsanguine Rhesus monkey model. The results showed that AGL could effectively remove alpha-Gal xenoantigens on pRBC membrane and reduce hemagglutination. The combination of mPEG modification with AGL treatment could significantly increased compatibility between pRBCs and Rhesus monkey sera. Modified pRBCs were detectable in Rhesus monkey blood at 12 hours after transfusion, and their survival time was 40 hours in the immunosuppressant-treated Rhesus monkey. In vivo survival rates of pRBCs were 38% in exsanguine Rhesus monkey at 8 hours after transfusion, and during that time, the hemoglobin and hematocrit of Rhesus monkey were maintained at the same level as before it lost blood. It is concluded that the modified pRBC can be safely transfused into Rhesus monkey and relieve the anemic symptom exsanguine Rhesus monkey. It suggested that pRBC can be hopefully used as a blood substitute for primate and human in the future.

Animals↗
PubMed2005

Nucleotide excision repair in chromatin and the right of entry.

DNA is packaged with histones and other accessory proteins into chromatin in eukaryotic cells. It is well established that the assembly of DNA into chromatin affects induction of DNA damage as well as repair of the damage. How the DNA repair machinery detects a lesion and 'fixes it' in chromatin has been an intriguing question since the dawn of understanding DNA packaging in chromatin. Direct recognition/binding by damaged DNA binding proteins is one obvious tactic to detect a lesion. Rearrangement of chromatin structure during DNA repair was reported more than two decades ago. This early observation suggests that unfolding of chromatin structure may be required to facilitate DNA repair after lesions are detected. Cells can also exploit DNA processing events to assist DNA repair. Transcription coupled repair (TCR) is such an example. During TCR, an RNA polymerase blocked by a lesion, may act as a signal to recruit DNA repair machinery. Possible roles of histone modification enzymes, ATP-dependent chromatin remodeling complexes and chromatin assembly factors in DNA repair are discussed.

Acetylation↗
PubMed2005

In vivo fragmentation of heparan sulfate by heparanase overexpression renders mice resistant to amyloid protein A amyloidosis.

Amyloid diseases encompass >20 medical disorders that include amyloid protein A (AA) amyloidosis, Alzheimer's disease, and type 2 diabetes. A common feature of these conditions is the selective organ deposition of disease-specific fibrillar proteins, along with the sulfated glycosaminoglycan, heparan sulfate. We have generated transgenic mice that overexpress human heparanase and have tested their susceptibility to amyloid induction. Drastic shortening of heparan sulfate chains was observed in heparanase-overproducing organs, such as liver and kidney. These sites selectively escaped amyloid deposition on experimental induction of inflammation-associated AA amyloidosis, as verified by lack of material staining with Congo Red, as well as lack of associated polysaccharide, whereas the same tissues from control animals were heavily infiltrated with amyloid. By contrast, the spleens of transgenic mice that failed to significantly overexpress heparanase contained heparan sulfate chains similar in size to those of control spleen and remained susceptible to amyloid deposition. Our findings provide direct in vivo evidence that heparan sulfate is essential for the development of amyloid disease.

Amyloidosis↗
PubMed2005

[Preparation of transfusable human universal red blood cell with recombinant alpha-galactosidase].

In order to meet the demand for safe transfusion in special conditions and to utilize the donated blood supply efficiently, technology has been developed to convert erythrocytes from type A, B, or AB to "universal donor" blood. Conversion of blood type B to O was performed by means of recombinant alpha-galactosidase digestion. The results showed that blood type B to O was converted successfully, 1 transfusion unit of red cells of group B (100 ml totally) could converted to universal blood cells in the optimal conditions including pH 5.6, 26 degrees C, 2 hours, obturation and sterilization. It is concluded that the universal red blood cells converted from group B to group O are conformed to demand of identification rules of biological products, no harmful effects of alpha-galactosidase on cell structure and function are observed. The converted red cells can stored in 4 degrees C for 21 days.

ABO Blood-Group System↗
PubMed2004

[Spectroscopic characterization of Tm3+ ions in zinc tungstate crystals].

A series of high quality ZnWO4:Tm3+ single crystals were grown with Czochralski method. The concentrations of thulium ions were 0.1, 0.3, 0.5, 0.7 and 1.0 mol%, respectively. When ZnWO4:Tm3+ single crystals were excited by 807 nm laser, the up-conversion luminescence peaks at 486 and 695 nm which corresponded to transitions 1G4-->3H6 and 3F3-->3 H6, respectively, were detected. The intensity of 695 nm luminescence varied with the pump power in a double logarithmic relation whose indexes were 0.8 and 1.1 orders, respectively.

Crystallization↗
PubMed2003

Processing of macromolecular heparin by heparanase.

Heparanase is an endo-glucuronidase expressed in a variety of tissues and cells that selectively cleaves extracellular and cell-surface heparan sulfate. Here we propose that this enzyme is involved also in the processing of serglycin heparin proteoglycan in mouse mast cells. In this process, newly synthesized heparin chains (60-100 kDa) are degraded to fragments (10-20 kDa) similar in size to commercially available heparin (Jacobsson, K. G., and Lindahl, U. (1987) Biochem. J. 246, 409-415). A fraction of these fragments contains the specific pentasaccharide sequence required for high affinity binding to antithrombin implicated with anticoagulant activity. Rat skin heparin, which escapes processing in vivo, was used as a substrate in reaction with recombinant human heparanase. An incubation product of commercial heparin size retained the specific pentasaccharide sequence, although oligosaccharides (3-4 kDa) containing this sequence could be degraded by the same enzyme. Commercial heparin was found to be a powerful inhibitor (I50 approximately 20 nM expressed as disaccharide unit, approximately 0.7 nM polysaccharide) of heparanase action toward antithrombin-binding oligosaccharides. Cells derived from a serglycin-processing mouse mastocytoma expressed a protein highly similar to other mammalian heparanases. These findings strongly suggest that the intracellular processing of the heparin proteoglycan polysaccharide chains is catalyzed by heparanase, which primarily cleaves target structures distinct from the antithrombin-binding sequence.

Acetylation↗
PubMed2003

Targeted disruption of a murine glucuronyl C5-epimerase gene results in heparan sulfate lacking L-iduronic acid and in neonatal lethality.

The glycosaminoglycan, heparan sulfate (HS), binds proteins to modulate signaling events in embryogenesis. All identified protein-binding HS epitopes contain l-iduronic acid (IdoA). We report that targeted disruption of the murine d-glucuronyl C5-epimerase gene results in a structurally altered HS lacking IdoA. The corresponding phenotype is lethal, with renal agenesis, lung defects, and skeletal malformations. Unexpectedly, major organ systems, including the brain, liver, gastrointestinal tract, skin, and heart, appeared normal. We find that IdoA units are essential for normal kidney, lung, and skeletal development, albeit with different requirement for 2-O-sulfation. By contrast, major early developmental events known to critically depend on heparan sulfate apparently proceed normally even in the absence of IdoA.

Animals↗
PubMed2003

A transcriptional pause synchronizes translation with transcription in the tryptophanase operon leader region.

Regulation of transcription of the tryptophanase operon requires that translation of its leader peptide coding region, tnaC, be coupled with its transcription. We show in vitro that a transcription pause site exists at the end of the tnaC coding region and that translation of tnaC releases the paused transcription complex, coupling transcription with translation.

Amino Acid Sequence↗
PubMed2002

Instruction of translating ribosome by nascent peptide.

Expression of the tryptophanase operon of Escherichia coli is regulated by catabolite repression and tryptophan-induced transcription antitermination. An induction site activated by l-tryptophan is created in the translating ribosome during synthesis of TnaC, the 24-residue leader peptide. Replacing the tnaC stop codon with a tryptophan codon allows tryptophan-charged tryptophan transfer RNA to substitute for tryptophan as inducer. This suggests that the ribosomal A site occupied by the tryptophanyl moiety of the charged transfer RNA is the site of induction. The location of tryptophan-12 of nascent TnaC in the peptide exit tunnel was crucial for induction. These results show that a nascent peptide sequence can influence translation continuation and termination within a translating ribosome.

Aminoglycosides↗
PubMed2002

Analysis of tryptophanase operon expression in vitro: accumulation of TnaC-peptidyl-tRNA in a release factor 2-depleted S-30 extract prevents Rho factor action, simulating induction.

Expression of the tryptophanase (tna) operon in Escherichia coli is regulated by catabolite repression and tryptophan-induced transcription antitermination. The key feature of this antitermination mechanism has been shown to be the retention of uncleaved TnaC-peptidyl-tRNA in the translating ribosome. This ribosome remains stalled at the tna stop codon and blocks the access of Rho factor to the tna transcript, thereby preventing transcription termination. In normal S-30 preparations, synthesis of a TnaC peptide containing arginine instead of tryptophan at position 12 (Arg(12)-TnaC) was shown to be insensitive to added tryptophan, i.e. Arg(12)-TnaC-peptidyl-tRNA was cleaved, and there was normal Rho-dependent transcription termination. When the S-30 extract used was depleted of release factor 2, Arg(12)-TnaC-tRNA(Pro) was accumulated in the absence or presence of added tryptophan. Under these conditions the accumulation of Arg(12)-TnaC-tRNA(Pro) prevented Rho-dependent transcription termination, mimicking normal induction. Using a minimal in vitro transcription system consisting of a tna template, RNA polymerase, and Rho, it was shown that RNA sequences immediately adjacent to the tnaC stop codon, the presumed boxA and rut sites, contributed most significantly to Rho-dependent termination. The tna boxA-like sequence appeared to serve as a segment of the Rho "entry" site, despite its likeness to the boxA element.

Arginine↗
PubMed2002

Identification and characterization of a mammalian 14-kDa phosphohistidine phosphatase.

Protein histidine phosphorylation in eukaryotes has been sparsely studied compared to protein serine/threonine and tyrosine phosphorylation. In an attempt to rectify this by probing porcine liver cytosol with the phosphohistidine-containing peptide succinyl-Ala-His(P)-Pro-Phe-p-nitroanilide (phosphopeptide I), we observed a phosphatase activity that was insensitive towards okadaic acid and EDTA. This suggested the existence of a phosphohistidine phosphatase different from protein phosphatase 1, 2A and 2C. A 1000-fold purification to apparent homogeneity gave a 14-kDa phosphatase with a specific activity of 3 micro mol.min-1.mg-1 at pH 7.5 with 7 micro m phosphopeptide I as substrate. Partial amino-acid sequence determination of the purified porcine enzyme by MS revealed similarity with a human sequence representing a human chromosome 9 gene of hitherto unknown function. Molecular cloning from a human embryonic kidney cell cDNA-library followed by expression and purification, yielded a protein with a molecular mass of 13 700 Da, and an EDTA-insensitive phosphohistidine phosphatase activity of 9 micro mol.min-1.mg-1 towards phosphopeptide I. No detectable activity was obtained towards a set of phosphoserine-, phosphothreonine-, and phosphotyrosine peptides. Northern blot analysis indicated that the human phosphohistidine phosphatase mRNA was present preferentially in heart and skeletal muscle. These results provide a new tool for studying eukaryotic histidine phosphorylation/dephosphorylation.

Amino Acid Sequence↗
PubMed1998

Cloning and Sequencing of Rhizobium huakuii exoA Gene Encoding a Glucosyl-transferase.

From exoR'-11 which could complement two Exo(-) mutants of R. huakuii 107: NA03 and NA10, a 2.0kb BglI fragment was subcloned in pRK415. The resulted plasmid pJB-H701 could restore the Exo(-) phenotype of NA03 and NA10. The complete nucleotide sequence of the fragment was determined, which contains the structural genes of a glucosyl-transferase, as well as the 5'- and 3'- flanking regions. An open reading frame of 984 base pairs was identified as R. huakuii exoA gene. The MW of ExoA, as deduced from the nucleotide sequence, was estimated as 35 kD. Comparison of the nucleotide sequences revealed a high similarity between exoA genes of R. huakuii and R. meliloti. The deduced amino acid sequence of R. huakuii ExoA also showed a high similarity with that of R. meliloti ExoA. Furthermore, the exoA-lacZ transcription fusion gene was constructed and the expression of exoA-lacZ was analyzed.

Journal Article↗