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Cloning and nucleotide sequence of the phosphoenolpyruvate carboxylase-coding gene of Corynebacterium glutamicum ATCC13032 总被引:5,自引:0,他引:5
As a first step in determining the importance of the anaplerotic function of phosphoenolpyruvate carboxylase (PEPC) in amino acid biosynthesis, the ppc gene coding for PEPC of Corynebacterium glutamicum ATCC13032 has been cloned by complementation of an Escherichia coli ppc mutant strain. PEPC activity encoded by the cloned gene is not affected by acetyl-CoA under conditions where the E. coli enzyme is strongly activated, whereas acetyl-CoA is able to relieve inhibition by L-aspartate used singly or in combination with alpha-ketoglutarate. Amplification of the ppc gene in a C. glutamicum lysine-excreting strain resulted in increased PEPC-specific activity and lysine productivity. The nucleotide sequence of a DNA fragment of 4885 bp encompassing the ppc gene has been determined. At the amino acid level, PEPC from C. glutamicum presents overall a high degree of similarity with corresponding enzymes from three different organisms. The location of some strictly conserved regions may have important implications for PEPC activity and allostery. 相似文献
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The aspartase gene (aspA) of Pseudomonas fluorescens was cloned and the nucleotide sequence of the 2,066-base-pair DNA fragment containing the aspA gene was determined. The amino acid sequence of the protein deduced from the nucleotide sequence was confirmed by N- and C-terminal sequence analysis of the purified enzyme protein. The deduced amino acid composition also fitted the previous amino acid analysis results well (Takagi et al. (1984) J. Biochem. 96, 545-552). These results indicate that aspartase of P. fluorescens consists of four identical subunits with a molecular weight of 50,859, composed of 472 amino acid residues. The coding sequence of the gene was preceded by a potential Shine-Dalgarno sequence and by a few promoter-like structures. Following the stop codon there was a structure which is reminiscent of the Escherichia coli rho-independent terminator. The G + C content of the coding sequence was found to be 62.3%. Inspection of the codon usage for the aspA gene revealed as high as 80.0% preference for G or C at the third codon position. The deduced amino acid sequence was 56.3% homologous with that of the enzyme of E. coli W (Takagi et al. (1985) Nucl. Acids Res. 13, 2063-2074). Cys-140 and Cys-430 of the E. coli enzyme, which had been assigned as functionally essential (Ida & Tokushige (1985) J. Biochem. 98, 793-797), were substituted by Ala-140 and Ala-431, respectively, in the P. fluorescens enzyme. 相似文献
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The nucleotide sequence of the glg B gene, coding for branching enzyme (EC 2.4.1.18), was elucidated. It consists of 2181 base pairs specifying a protein of 727 amino acids. The deduced amino acid sequence was consistent with the amino acid analysis that was obtained with the pure protein as well as with the molecular weight determined from sodium dodecyl sulfate-gel electrophoresis. The deduced amino acid sequence was also consistent with the amino-terminal amino acid sequence and the amino acid sequence analysis of various peptides obtained from CNBr degradation of purified branching enzyme. 相似文献
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Molecular cloning and nucleotide sequence of the beta-lytic protease gene from Achromobacter lyticus. 总被引:3,自引:2,他引:1
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Two bacteriolytic enzymes secreted by Achromobacter lyticus M497-1 were purified and identified as being very similar (considering their amino acid composition and N-terminal sequence) to alpha- and beta-lytic proteases from Lysobacter enzymogenes. A 1.8-kb EcoRI fragment containing the structural gene for beta-lytic protease was cloned from A. lyticus chromosomal DNA. The protein sequence deduced from the nucleotide sequence was identical to the known sequence of beta-lytic protease, except for six residues. The nucleotide sequence revealed that the mature enzyme is composed of 179 amino acid residues with an additional 195 amino acids at the amino-terminal end of the enzyme, which includes the signal peptide, thus indicating that the enzyme is synthesized as a precursor protein. 相似文献
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Branched-chain amino acid aminotransferase of Escherichia coli: nucleotide sequence of the ilvE gene and the deduced amino acid sequence 总被引:6,自引:0,他引:6
The ilvE gene of the Escherichia coli K-12 ilvGEDA operon, which encodes branched-chain amino acid aminotransferase [EC 2.6.1.42], was cloned. The nucleotide sequence of 1.5 kilobase pairs containing the gene was determined. The coding region of the ilvE gene contained 927 nucleotide residues and could encode 309 amino acid residues. The predicted molecular weight, amino acid composition and the sequence of the N-terminal 15 residues agreed with the enzyme data reported previously (Lee-Peng, F.-C., et al. (1979) J. Bacteriol. 139, 339-345). From the deduced amino acid sequence, the secondary structure was predicted. 相似文献
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Nucleotide sequence of the murE gene of Escherichia coli 总被引:1,自引:0,他引:1
The nucleotide sequence of the murE gene encoding the diaminopimelic acid adding enzyme of Escherichia coli is reported. The coding region consisted of 1413 base pairs and was separated from the ftsI (penicillin-binding protein 3) gene by 61 base pairs. The deduced primary structure of MurE comprised 471 amino acid residues with a molecular mass of 50.6 kilodaltons. 相似文献
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The nucleotide sequence of the G6-amylase gene from alkalophilic Bacillus sp. H-167 was determined. The open reading frame of the gene consisted of 2865 base pairs, encoding 955 amino acids. The NH2-terminal amino acid sequence analysis of the G6-amylase indicated that the enzyme had a single peptide of 33 amino acid residues and the mature enzyme was composed of 922 amino acids, giving a molecular mass of 102,598. Identity of the NH2-terminal amino acid sequences among each component of the multiform G6-amylase suggested the proteolytic processing of the COOH-terminal side of the enzyme. The DNA sequence and the deduced amino acid sequence of the G6-amylase gene showed no homology with those of other bacterial alpha-amylases although the consensus amino acid sequences of the active center were well conserved. 相似文献
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The complete nucleotide sequence of the methotrexate-resistant dihydrofolate reductase (DHFR) gene borne by the plasmid R67 was determined. The gene is 234 bp long and codes for 78 amino acids. The polypeptide deduced from the DNA sequence is in perfect agreement with the previously published amino acid sequence. Comparison of the nucleotide sequence with the one determined for the R388-encoded DHFR indicates that 75% of the nucleotides are conserved in the two genes. The 3′ end of the R67 gene can be modified without altering significantly the activity of the enzyme. 相似文献
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Nucleotide sequence of the gene for cholesterol oxidase from a Streptomyces sp. 总被引:9,自引:3,他引:6
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The nucleotide sequence of a 2.1-kilobase-pair fragment containing the Streptomyces choA gene, which codes a secreted cholesterol oxidase, was determined. A single open reading frame encodes a mature cholesterol oxidase of 504 amino acids, with a calculated Mr of 54,913. The leader peptides extend over 42 amino acids and have the characteristics of a signal sequence, including basic amino acids near the amino terminus and a hydrophobic core near the signal cleavage site. Analyses of the total amino acid composition and amino acid sequencing of the first 21 amino acids from the N terminus of the purified extracellular enzyme agree with the values deduced from nucleotide sequencing data. 相似文献
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《Journal of Fermentation and Bioengineering》1994,77(3):315-319
We found that the structural gene for monoamine oxidase was located at 30.9 min on the Escherichia coli chromosome. Deletion analysis showed that two amine oxidase genes are located in this region. The nucleotide sequence of one of the two genes was determined. The peptide sequence of the first 40 amino acids from the N terminus of monoamine oxidase purified from E. coli agrees with that deduced from the nucleotide sequence of the gene. The leader peptide extends over 30 amino acids. The nucleotide sequence of the gene and amino acid sequence of the predicted mature enzyme (M.W. 81,295) were highly homologous to those of the maoAK gene and monoamine oxidase from Klebsiella aerogenes, respectively. From these results and analysis of the enzyme activity, we concluded that the gene encodes for monoamine oxidase (maoAE). The tyrosyl residue, which may be converted to topa quinone in the E. coli enzyme, was located by comparison with amino acid sequences at the cofactor sites in other copper/topa quinone-containing amine oxidases. 相似文献
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Molecular cloning and nucleotide sequence of the gene encoding an endo-1,4-beta-glucanase from Bacillus sp. KSM-330. 总被引:1,自引:0,他引:1
The gene encoding an acid endo-1,4-beta-glucanase from Bacillus sp. KSM-330 was cloned into the HindIII site of pBR322 and expressed in Escherichia coli HB101. The recombinant plasmid contained a 3.1 kb HindIII insert, 1.8 kb of which was sufficient for the expression of endoglucanase activity in E. coli HB101. Nucleotide sequencing of this region (1816 bp) revealed an open reading frame of 1389 bp. The protein deduced from this sequence was composed of 463 amino acids with an Mr of 51882. The deduced amino acid sequence from amino acids 56 through 75 coincided with the amino-terminal sequence of the endoglucanase, Endo-K, purified from culture of Bacillus sp. KSM-330. The deduced amino acid sequence of Endo-K had 30% homology with that of the celA enzyme from Clostridium thermocellum NCIB 10682 and 25% homology with that of the enzyme from Cellulomonas uda CB4. However, the Endo-K protein exhibited no homology with respect to either the nucleotide or the amino acid sequences of other endoglucanases from Bacillus that had been previously characterized. These results indicate that the gene for Endo-K in Bacillus sp. KSM-330 has evolved from an ancestral gene distinct from that of other Bacillus endoglucanases. 相似文献
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Nucleotide sequence of the beta-cyclodextrin glucanotransferase gene of alkalophilic Bacillus sp. strain 1011 and similarity of its amino acid sequence to those of alpha-amylases. 总被引:5,自引:2,他引:3
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The nucleotide sequence of the gene for cyclodextrin glucanotransferase of alkalophilic Bacillus sp. strain 1011 was determined. The deduced amino acid sequence at the NH2-terminal side of the enzyme showed a high homology with the sequences of alpha-amylase in the three regions which constitutes the active centers of alpha-amylases. 相似文献
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R E Perkins S C Conroy B Dunbar L A Fothergill M F Tuite M J Dobson S M Kingsman A J Kingsman 《The Biochemical journal》1983,211(1):199-218
The complete amino acid sequence of yeast phosphoglycerate kinase, comprising 415 residues, was determined. The sequence of residues 1-173 was deduced mainly from nucleotide sequence analysis of a series of overlapping fragments derived from the relevant portion of a 2.95-kilobase endonuclease-HindIII-digest fragment containing the yeast phosphoglycerate kinase gene. The sequence of residues 174-415 was deduced mainly from amino acid sequence analysis of three CNBr-cleavage fragments, and from peptides derived from these fragments after digestion by a number of proteolytic enzymes. Cleavage at the two tryptophan residues with o-iodosobenzoic acid was also used to isolate fragments suitable for amino acid sequence analysis. Determination of the complete sequence now allows a detailed interpretation of the existing high-resolution X-ray-crystallographic structure. The sequence -Ile-Ile-Gly-Gly-Gly- occurs twice in distant parts of the linear sequence (residues 232-236 and 367-371). Both these regions contribute to the nucleoside phosphate-binding site. A comparison of the sequence of yeast phosphoglycerate kinase reported here with the sequences of phosphoglycerate kinase from horse muscle and human erythrocytes shows that the yeast enzyme is 64% identical with the mammalian enzymes. The yeast has strikingly fewer methionine, cysteine and tryptophan residues. 相似文献
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Entire nucleotide sequence of the pullulanase gene of Klebsiella aerogenes W70. 总被引:10,自引:9,他引:10
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We determined the entire nucleotide sequence of the Klebsiella aerogenes W70 pullulanase gene (pulA) contained on a 4.2-kilobase-pair fragment of plasmid pPB174. The amino acid composition deduced from an open reading frame of 3,288 base pairs agreed closely with that determined for the intracellular pullalanase. The precursor enzyme consisted of 1,096 amino acid residues and contained a hydrophobic N-terminal signal peptide and the consensus sequence for the bacterial prelipoprotein signal peptide cleavage site. 相似文献
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The nucleotide sequence of the G6 -amylase gene from alkalophilic Bacillus sp. H-167 was determined. The open reading frame of the gene consisted of 2865 base pairs, encoding 955 amino acids. The NH2 -terminal amino acid sequence analysis of the G6 -amylase indicated that the enzyme had a single peptide of 33 amino acid residues and the mature enzyme was composed of 922 amino acids, giving a molecular mass of 102598. Identity of the NH2 -terminal amino acid sequences among each component of the multiform G6 -amylase suggested the proteolytic processing of the COOH-terminal side of the enzyme. The DNA sequence and the deduced amino acid sequence of the G6 -amylase gene showed no homology with those of other bacterial α-amylases although the consensus amino acid sequences of the active center were well conserved. 相似文献
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Nucleotide sequence of Escherichia coli purF and deduced amino acid sequence of glutamine phosphoribosylpyrophosphate amidotransferase 总被引:26,自引:0,他引:26
J Y Tso H Zalkin M van Cleemput C Yanofsky J M Smith 《The Journal of biological chemistry》1982,257(7):3525-3531
The Escherichia coli gene purF, coding for 5-phosphoribosylamine:glutamine pyrophosphate phosphoribosyltransferase (amidophosphoribosyltransferase) was subcloned from a ColE1-purF plasmid into pBR322. Amidophosphoribosyltransferase levels were elevated more than 5-fold in the ColE1-purF plasmid-bearing strain compared to the wild type control, and a further 10- to 13-fold elevation was observed in several pBR322 derivatives. The nucleotide sequence of a 2478-base pair PvuI-HinfI fragment encoding purF was determined. The purF45 structural gene codes for a 56,395 Mr protein chain having 504 amino acid residues. Methionine-1 is removed by processing in vivo leaving cysteine as the NH2-terminal residue. The deduced amino acid sequence was confirmed by comparisons with the NH2-terminal amino acid sequence determined by automated Edman degradation (Tso, J. Y., Hermodson, M. A., and Zalkin, H. (1982) J. Biol. Chem. 257, 3532-3536) and amino acid analyses of CNBr peptides including a 4-residue peptide from the CO2H terminus of the enzyme. Nucleotide sequences characteristic of bacterial promoter-operator regions were identified in the 5' flanking region. The coding region appears to be preceded by a 277-297 nucleotide mRNA leader. A deletion removing the putative promoter-operator region results in defective purF expression. 相似文献