Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.7 (DNA polymerase)
 17,007 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An enconuclease activity that reacts with x-irradiated DNA is present in extracts of E. coli. By using centrifugal methods to monitor the conversion of the supercoiled, circular double-stranded DNA for phage phi-x-174 (replicative form) or PM2 to the relaxed circular form it was possible to quantitate the rate of radiation induced endonuclease-sensitive sites in the DNA. For every single-strand break induced by x-rays under aerobic irradiation conditions, there is approximately one induced site sensitive to this endonuclease activity. Under irradiation conditions (addition OF Potassium iodide) that dramatically reduce rates of single-strand breaks and "alkalilabile" lesions, the number of endonuclease-sensitive sites relative to single-strand breaks increase approximatley 4-fold. This nuclease is present in several strains of E. coli B and K12, including mutants deficient in DNA polymerase I, recombination gene products (rec mutants), ultraviolet light incision enzyme (uvr A mutant), and endonuclease II. It is suggested that this endonuclease may be involved in an excision repair process for damages incurred in DNA by ionizing radiation.
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PMID:Endonucleolytic incision of x-irradiated deoxyribonucleic acid by extracts of Escherichia coli. 109 50

Insulin resistance is an early predictor of development of noninsulin-dependent diabetes mellitus (NIDDM) in Pima Indians, a population with the highest reported prevalence of NIDDM. The insulin receptor plays a central role in mediating insulin action, and previous studies have demonstrated that mutations in the insulin receptor gene may cause insulin resistance. Therefore, we have cloned the insulin receptor cDNA from an insulin-resistant Pima Indian to determine if there is a mutation in the patient's insulin receptor gene. We obtained nine cDNA clones spanning exons 4-10 and 12-22 of the patient's insulin receptor gene. Polymorphisms in the nucleotide sequences for codons 523 (Ala), 1058 (His), and 1062 (Leu) provided useful markers to differentiate the patient's two alleles of the insulin receptor gene. These substitutions were silent, in that they did not alter the predicted amino acid sequence. The sequence of exons 1-3 and 11 was determined directly from genomic DNA that had been amplified using the polymerase chain reaction catalyzed by Taq DNA polymerase. Other investigators have reported defects in insulin binding and insulin receptor tyrosine kinase activity in diabetic Pima Indians. However, we did not detect any mutations in this patient's insulin receptor gene. Thus, these observations are consistent with the interpretation that the defects in insulin receptor function are acquired rather than derived from defects in the primary structure of the receptor.
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PMID:The amino acid sequence of the insulin receptor is normal in an insulin-resistant Pima Indian. 231 37

Using the technique of ultraviolet-mediated cross-linking of substrate deoxynucleoside triphosphates (dNTPs) to their acceptor site [Abraham, K. I., & Modak, M. J. (1984) Biochemistry 23, 1176-1182], we have labeled the Klenow fragment of Escherichia coli DNA polymerase I (Pol I) with [alpha-32P]dTTP. Covalent cross-linking of [alpha-32P]dTTP to the Klenow fragment is shown to be at the substrate binding site by the following criteria: (a) the cross-linking reaction requires dTTP in its metal chelate form; (b) dTTP is readily competed out by other dNTPs as well as by substrate binding site directed reagents; (c) labeling with dTTP occurs at a single site as judged by peptide mapping. Under optimal conditions, a modification of approximately 20% of the enzyme was achieved. Following tryptic digestion of the [alpha-32P]dTTP-labeled Klenow fragment, reverse-phase high-performance liquid chromatography demonstrated that 80% of the radioactivity was contained within a single peptide. The amino acid composition and sequence of this peptide identified it as the peptide spanning amino acid residues 876-890 in the primary sequence of E. coli Pol I. Chymotrypsin and Staphylococcus aureus V8 protease digestion of the labeled tryptic peptide in each case yielded a single smaller fragment that was radioactive. Amino acid analysis and sequencing of these smaller peptides further narrowed the dTTP cross-linking site to within the region spanning residues 876-883. We concluded that histidine-881 is the primary attachment site for dTTP in E. coli DNA Pol I, since during amino acid sequencing analysis of all three radioactive peptides loss of the histidine residue at the expected cycle is observed.
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PMID:Photoaffinity labeling of the thymidine triphosphate binding domain in Escherichia coli DNA polymerase I: identification of histidine-881 as the site of cross-linking. 332 6