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)

Intact RNA from various rat organs was isolated by an efficient and rapid method. This method of RNA isolation is a modification of an earlier method that uses guanidinium isothiocynate followed by extraction in the presence of sarcosyl, acetate and phenol. The RNA obtained by the method reported here was comparable with the RNA prepared by the CsCl2 ultracentrifugation method and the commercially available kit based on published methods. The quality of RNA was found suitable for Northern blotting analysis, RNase protection assays and reverse transcriptase-polymerase chain reaction (RT-PCR). Since reverse transcriptase is active in the buffer used for Taq DNA polymerase, only one reaction needs to be set up. We also found that the use of aurintricarboxylic acid in the RNA preparation prevents the degradation of RNA during storage. Expression of low density lipoprotein (LDL) receptor, apolipoprotein (apo) AI, AII and AIV mRNAs were quantified in various rat organs. Our results indicated that rat LDL receptor mRNA is expressed in several organs whereas apoAI and AIV mRNAs were expressed mainly in the liver and intestine. However, apo AII mRNA is expressed mainly in the liver. Unlike mice and some species of monkeys, in the rat apoAI mRNA is expressed at 5-6 times higher levels in the intestine compared to liver. Apo AIV mRNA abundance was also found to be several fold higher in intestine compared to hepatic tissues. We present here, for the first time, data on the absolute amounts of LDL receptor, apoAI, AII and AIV mRNA in various rat organs which were quantified by a novel RNase protection/solution hybridization assay.
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PMID:Expression of low density lipoprotein receptor, apolipoprotein AI, AII and AIV in various rat organs utilizing an efficient and rapid method for RNA isolation. 137 76

Small protein domains, capable of specific binding to different target proteins have been selected using combinatorial approaches. These binding proteins, called affibodies, were designed by randomization of 13 solvent-accessible surface residues of a stable alpha-helical bacterial receptor domain Z, derived from staphylococcal protein A. Repertoires of mutant Z domain genes were assembled and inserted into a phagemid vector adapted for monovalent phage display. Two libraries, each comprising approximately 4 x 10(7) transformants, were constructed using either an NN(G/T) or an alternative (C/A/G)NN degeneracy. Biopanning against the target proteins Taq DNA polymerase, human insulin, and a human apolipoprotein A-1 variant, showed that in all cases significant enrichments were obtained by the selection procedures. Selected clones were subsequently expressed in Escherichia coli and analyzed by SDS-PAGE, circular dichroism spectroscopy, and binding studies to their respective targets by biospecific interaction analysis. The affibodies have a secondary structure similar to the native Z domain and have micromolar dissociation constants (KD) for their respective targets.
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PMID:Binding proteins selected from combinatorial libraries of an alpha-helical bacterial receptor domain. 925 93

Several men were examined for association between restriction fragment length polymorphism (RELP) Xba I (exon 26) number of tandem repeats in 3'-hypervariable region of the apolipoprotein-B gene and serum levels of cholesterol and triglyceride. These two types of polymorphism were studied. An association of the Xba I site and alleles containing more repeats in the 3'-hypervariable region with higher cholesterol and triglyceride was observed. 32 patients with CHD aged 24-56 years were examined. All the patients are males with clinical picture of CHD (stable angina pectoris of II-III functional classes) and dyslipoproteinemia of II a, II b and IV types by D. S. Fredrickson. Xba-I polymorphism of apo-B gene was detected by DNA polymerase reaction method. The following Xba-I genotypes were distinguished: X1X1 (absence of Xba I site); X1X2 (heterozygosity on Xba I site) and X2X2 (homozygosity on Xba-I site). Lipantil (fenofibratte) was prescribed in a dose of 300 mg daily after meals (100 mg three times a day). Data obtained show that DNA polymorphism of apo-B gene not only influences plasma lipids concentration but also determines effectiveness of hypolipidemic therapy. Hypolipidemic effect of lipantil depends on Xba-I site presents in apo-B gene and is significantly expressed in homozygous patients with X1X1 genotype.
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PMID:[The apoB genotype and the efficacy of hypolipidemic therapy]. 969 63

Here we show that robust and small protein ligands can be used for affinity capture of recombinant proteins from crude cell lysates. Two ligands selectively binding to bacterial Taq DNA polymerase and human apolipoprotein A-1(M), respectively, were used in the study. The ligands were selected from libraries of a randomized alpha-helical bacterial receptor domain derived from staphylococcal protein A and have dissociation constants in the micromolar range, which is typical after primary selection from these libraries consisting of approximately 40 million different members each. Using these ligands in affinity chromatography, both target proteins were efficiently recovered from crude cell lysates with high selectivities. No loss of column capacity or selectivity was observed for repeated cycles of sample loading, washing and low pH elution. Interestingly, column sanitation could be performed using 0. 5 M sodium hydroxide without significant loss of ligand performance. The results suggest that combinatorial approaches using robust protein domains as scaffolds can be a general tool in the process of designing purification strategies for biomolecules.
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PMID:Ligands selected from combinatorial libraries of protein A for use in affinity capture of apolipoprotein A-1M and taq DNA polymerase. 1086 85