Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.10.1 (ERK)
 95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Calcium 2-keto-L-gulonate (Ca-2-KLG, a key intermediate in vitamin C synthesis) is produced from calcium 2,5-diketo D-gluconate (Ca-2,5-DKG) by a variety of bacteria. A few bacterial species which efficiently convert glucose to Ca-2,5-DKG have been isolated in our laboratory. Our bacterial collection included species that possess the genes for production of Ca-2-KLG from Ca-2,5-DKG; however, the yield of the former is poor. A procedure for the preparation of spheroplasts in Ca-2,5-DKG- and Ca-2-KLG-producing bacteria was developed for the construction of recombinants (fusants), combining the genes for conversion of glucose to Ca-2-KLG efficiently by protoplast fusion. The standard procedure for spheroplast formation in Gram negative bacteria by the Tris-sucrose-EDTA-lysozyme system did not work in the organisms under investigation. The need for an alternative method was necessary. Our results show that, while the Tris-NaCl-EDTA-lysozyme system (pH 8.3) worked very well with bacterial strains of Gluconobacter oxydans (ATCC9937) and Acetobacter melanogenus (NCIM2259), the Tris-sucrose-EDTA-lysozyme system worked well for Erwinia herbicola (ATCC21998), Pseudomonas chlororaphis (NCIM2041) and Corynebacterium species (ATCC31090). However, none of these systems produced spheroplasts in Brevibacterium ketosoreductum (ATCC21914), for which a separate system is under development.
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PMID:A fast spheroplast formation procedure in some 2,5-diketo-D-gluconate- and 2-keto-L-gulonate- producing bacteria. 251 94

The quantitative analysis of 2-keto-L-gulonic acid (2-KLG) produced by microbial fermentation is described. 2-KLG is separated from other aldonic and ketoaldonic acids by high-performance liquid chromatography on an Aminex anion exchange column with ammonium formate or potassium phosphate as the eluant. This is a rapid and simple method for routine analysis of a large number of samples generated by fermentation studies. Gas chromatography--mass spectrometry permits the qualitative and quantitative analysis of nanogram levels of 2-keto-L-gulonate in complex media and provides confirmation of the HPLC results. The methodologies presented are useful for the analysis of a number of aldonic and ketoaldonic acids.
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PMID:Determination of 2-keto-L-gulonic and other ketoaldonic and aldonic acids produced by ketogenic bacterial fermentation. 377 40

Complementary DNA (cDNA) encoding a novel member of the receptor tyrosine kinase (RTK) family has been isolated from colon carcinoma tissue. Colon carcinoma kinase 4 (CCK-4) mRNA is highly expressed in human lung tissue and at lower levels in the thyroid gland and ovary. While no mRNA was found in human adult colon tissues, expression varied remarkably in colon carcinoma-derived cell lines. CCK-4 cDNA encodes a chicken KLG-related, 1071 amino acid-long transmembrane glycoprotein containing several genetic alterations within the RTK consensus sequences. These define CCK-4 as a catalytically inactive member of the RTK family of proteins and, in analogy to HER3, suggest a potentially tumor-characteristic role as a signal amplifier or modulator for an as yet unidentified kinase-competent partner.
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PMID:Colon carcinoma kinase-4 defines a new subclass of the receptor tyrosine kinase family. 747 40

A 220-bp fragment of PTK7 cDNA was previously cloned from normal human melanocyte RNAs by means of the reverse transcription-polymerase chain reaction [Lee, S.-T., Strunk, K.M., and Spritz, R.A. (1993) Oncogene 8, 3403-3410]. We now report the cloning of the human full-length PTK7 cDNA and its characterization. The 1,070 amino acid PTK7 polypeptide deduced from the cDNA sequence constitutes receptor protein tyrosine kinase (RPTK), but has several unusual residues in some of the highly conserved tyrosine kinase motifs. PTK7 mRNA was expressed at the highest level in a human erythroleukemia cell line among tested samples, and at relatively high levels in liver, lung, pancreas, kidney, placenta, and melanocytes. Human PTK7 is 72% identical to chick KLG, suggesting that PTK7 is homologous or possibly orthologous to chick KLG, and that these represent a new subfamily of RPTKs.
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PMID:Characterization of the human full-length PTK7 cDNA encoding a receptor protein tyrosine kinase-like molecule closely related to chick KLG. 888 11

Substrate selectivity of Gluconobacter oxydans (ATCC 9937) for 2,5-diketo-D-gluconic acid (2,5-DKG) production was investigated with glucose, gluconic acid, and gluconolactone in different concentrations using a resting-cell system. The results show that gluconic acid was utilized favorably by G. oxydans as substrate to produce 2,5-DKG. The strain was coupled with glucose dehydrogenase (GDH) and 2,5-DKG reductase for synthesis of 2-keto-L-gulonic acid (2-KLG), a direct precursor of L-ascorbic acid, from glucose. NADP and NADPH were regenerated between GDH and 2,5-DKG reductase. The mole yield of 2-KLG of this multienzyme system was 16.8%. There are three advantages for using the resting cells of G. oxydans to connect GDH with 2,5-DKG reductase for production of 2-KLG: gluconate produced by GDH may immediately be transformed into 2,5-DKG so that a series of problems generally caused by the accumulation of gluconate would be avoided; 2,5-DKG is supplied directly and continuously for 2,5-DKG reductase, so it is unnecessary to take special measures to deal with this unstable substrate as it was in Sonoyama's tandem fermentation process; and NADP(H) was regenerated within the system without any other components or systems.
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PMID:Substrate selectivity of Gluconobacter oxydans for production of 2,5-diketo-D-gluconic acid and synthesis of 2-keto-L-gulonic acid in a multienzyme system. 1156 24

We have identified nine oligopeptide transporter (OPT) orthologs (AtOPT1 to AtOPT9) in Arabidopsis. These proteins show significant sequence similarity to OPTs of Candida albicans (CaOpt1p), Schizosaccharomyces pombe (Isp4p), and Saccharomyces cerevisiae (Opt1p and Opt2p). Hydrophilicity plots of the OPTs suggest that they are integral membrane proteins with 12 to 14 transmembrane domains. Sequence comparisons showed that the AtOPTs form a distinct subfamily when compared with the fungal OPTs. Two highly conserved motifs (NPG and KIPPR) were found among all OPT members. The identification of multiple OPTs in Arabidopsis suggests that they may play different functional roles. This idea is supported by the fact that AtOPTs have a distinct, tissue-specific expression pattern. The cDNAs encoding seven of the AtOPTs were cloned into a yeast vector under the control of a constitutive promoter. AtOPT4 expressed in S. cerevisiae mediated the uptake of KLG-[3H]L. Similarly, expression of five of the seven AtOPT proteins expressed in yeast conferred the ability to uptake tetra- and pentapeptides as measured by growth. This study provides new evidence for multiple peptide transporter systems in Arabidopsis, suggesting an important physiological role for small peptides in plants.
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PMID:An oligopeptide transporter gene family in Arabidopsis. 1178 49

A screening method has been developed to support randomized mutagenesis of amino acids in the cofactor-binding pocket of the NADPH-dependent 2,5-diketo-D-gluconic acid (2,5-DKG) reductase. Such an approach could enable the isolation of an enzyme that can better catalyze the reduction of 2,5-DKG to 2-keto-L-gulonic acid (2-KLG) using NADH as a cofactor. 2-KLG is a valuable precursor to ascorbic acid, or vitamin C, and an enzyme with increased activity with NADH may be able to improve two potential vitamin C production processes. Previously we have identified three amino acid residues that can be mutated to improve activity with NADH as a cofactor. As a pilot study to show feasibility, a library was made with these three amino acids randomized, and 300 random colonies were screened for increased NADH activity. The activities of seven mutants with apparent improvements were verified using activity-stained native gels, and sequencing showed that the amino acids obtained were similar to some of those already discovered using rational design. The four most active mutants were purified and kinetically characterized. All of the new mutations resulted in apparent kcat values that were equal to or higher than that of the best mutant obtained through rational design. At saturating levels of cofactor, the best mutant obtained was almost twice as active with NADH as a cofactor as the wild-type enzyme is with NADPH. This screen is a valuable tool for improving 2,5-DKG reductase, and it could easily be modified for improving other aspects of this protein or similar enzymes.
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PMID:Verification of a novel NADH-binding motif: combinatorial mutagenesis of three amino acids in the cofactor-binding pocket of Corynebacterium 2,5-diketo-D-gluconic acid reductase. 1248 21

Based on the reported gene sequences, the segments containing 2-keto aldose reductase (2-KRA and B) genes were amplified by PCR from the plasmids and Erwinia sp. SCB125 each for gene expression and gene knocking out. Then cloning them into expression vector pBL and successfully expressing them with high enzyme activity in E. coli DH5 alpha. After their enzyme activities were proved, the work on gene knock out followed. Introducing the knock-out vector which distribute unstably during the cell division to the host strains Erwinia sp. SCB125. Screening firstly by the positive marker, one resistance which resulted from the expression of the resistance gene inserting inside the reductase genes and the negative marker, another resistance which outside the reductase genes in the vector. The strains selected out will be tested by further study. This work was the bases of blocking the pathway metabolism and constructing a recombinant strain that can produce 2-KLG directly from D-glucose by one-step fermentation.
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PMID:[Studies on gene knocking out of 2-keto aldose reductases from Erwinia sp. SCB125]. 1254 57

Two different 2,5-DKG reductases was purified from Corynebacterium sp. SCB3058, then its genomic DNA was used as template, a segment containing 2,5-DKG reductase I was amplified by PCR and cloned into pGEM3zf(+) to obtain the recombinant plasmid pGEM813. The sequence analysis showed that the cloned segment was 1107 bp in length, contained a single open reading frame of 834 nucleotides, which encoded a 34 kD protein consisted of 278 amino acids. After the primary control sequence was deleted, the expression vector pBL4 was constructed by plasmid pBL. With induction of temperature, a 34 kD protein was specifically expressed in E. coli DH5 alpha in high level. The expressed recombinant protein accounted for 20% of the total cell protein and had high specific enzyme activity. In spite of 30 degrees C, 37 degrees C or 42 degrees C induction, the specific activity of enzyme was almost the same level. These results indicate that most of the recombinant protein induced at 42 degrees C was existed with inclusion bodies. This work was the bases of constructing a recombinant Erwinia which can produce 2-KLG directly from D-glucose by one-step fermentation.
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PMID:[Cloning and expression in E. coli of 2,5-DKG reductase I from Corynebacterium]. 1254 22

Gluconobacter oxydans SCB329 only produce a little amount of 2-Keto-L-gulonic acid(2-KLG) from D-Sorbitol when growing alone; while Gluconobacter sp. SCB110 can transform D-Sorbitol to L-Sorbose and can not produce 2-KLG. 2-Keto-L-gulonic acid, the precursor of L-Ascorbic acid (Vitamin C) synthesis, was prepared directly with a high efficiency from D-Sorbitol by mixed culture of microorganism, which comprised Gluconobacter sp. SCB110 and Gluconobacter oxydans SCB329. The fermentation product from the mixed culture broth in the D-Sorbotol-containing medium was identified as 2-Keto-L-gulonic acid by HPLC, elementary analysis and infra-red adsorption spectrum.
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PMID:[Production of vitamin C precursor--2-keto-L-gulonic acid from D-sorbitol by mixed culture of microorganisms]. 1255 28


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