Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.5.1.3 (dihydrofolate reductase)
 5,819 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A new series of double-selection plasmids containing recombinant genes expressing the neomycin phosphotransferase (NEO) of transposon Tn5 and mouse dihydrofolate reductase (DHFR) in mammalian cells is described. Activity of the recombinant DHFR gene varied more than 50-fold, depending on the location of the simian virus 40 72 base-pair repeat or enhancer, which is part of the promoter of the NEO unit. A NEO-DHFR module with the enhancer located at the 3' end of the DHFR gene was inserted into a plasmid containing four tandem head-to-tail copies of the hepatitis B virus (HBV) genome and the new plasmid was used to transform DHFR- Chinese hamster ovary cells. In one of the cell lines obtained, an unrearranged copy of the HBV tetramer could be amplified 300-fold by increasing selective pressure with methotrexate, resulting in a proportional increase of the synthesis of HBV surface antigen. Four different mRNAs detected in the amplified cell line probably encode HBV core protein, pre-S and surface antigens, and the X protein. As a result of the DNA amplification, synthesis of HBV proteins is no longer restricted to resting cells. Integrated plasmid sequences appear to be stable during the amplification process.
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PMID:A recombinant Chinese hamster ovary cell line containing a 300-fold amplified tetramer of the hepatitis B genome together with a double selection marker expresses high levels of viral protein. 302 26

The small dermatan sulphate protein decorin interacts via its core protein with fibrillar collagens, and its glycosaminoglycan chains were proposed to be capable of self-association. It was therefore of interest to study the role of decorin in the contraction of cell-populated collagen lattices. Stable transfection of dihydrofolate reductase-deficient CHO cells with decorin cDNA resulted in impaired collagen lattice contraction. Using normal human skin fibroblasts in serum-free cultures, inclusion of 0.3 microM decorin in the culture medium also led to a delayed collagen gel contraction. Protein-free dermatan sulphate and the dermatan sulphate-degrading enzyme chondroitin ABC lyase were ineffective. Potential interactions between dermatan sulphate chains were studied by gel filtration. A shift in the elution position of [35S]sulphate-labelled decorin-derived glycosaminoglycans by unlabelled decorin could be observed only when the chains were prepared by trypsin. Chains liberated by beta-elimination or by cathepsin C were eluted at identical positions in the presence or absence of decorin. It is therefore unlikely, that the effect of decorin on collagen-gel retraction is brought about solely by glycosaminoglycan-glycosaminoglycan interactions.
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PMID:Modulation of collagen gel contraction by decorin. 866 Feb 78

The expression and processing of hepatitis C virus core protein was analyzed. Two protein bands, 21 kDa (P21), corresponding to the full-length core, and 19 kDa (P19), were detected as major products when core protein was expressed in the standard rabbit reticulocyte lysate system or in Sf9 insect cells. Core proteins with amino-terminal hexa-histidine tags were expressed which allowed the purification of the hexa-histidine P19 core with NI(2+)-NTA columns. With the help of mass spectrometry, the molecular weight of hexa-histidine-P19 was analyzed and its carboxy-terminus could be calculated. Fusion proteins of truncated core/core-E1 species fused to mouse dihydrofolate reductase (mDHFR) showed cleavage in the expected region. Cleavage sites could be determined by amino-terminal protein sequencing of the DHFR-fusion partner. Our data show that there are not one but two core products with an apparent molecular weight of about 19 kDa, ending either at amino acid leucine 179 or leucine 182, respectively. These cleavages in the hydrophobic, carboxy-terminal region of HCV core suggest processing by (a) recently proposed eucaryotic signal peptide peptidase(s) (F. Lyko et al. (1995) J. Biol. Chem. 270, 19873-19878). Furthermore, our results demonstrate that cleavage at these sites and the formation of the P19 species does not require previous processing at the signalase site (position 191/192) of the HCV-polyprotein.
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PMID:Hepatitis C virus core protein: carboxy-terminal boundaries of two processed species suggest cleavage by a signal peptide peptidase. 886 3

Recombinant and synthetic peptides derived from the human immunodeficiency virus type I (HIV-1) genome corresponding to portions of the envelope (env) and internal core protein (gag) were examined for their immunoregulatory effects on the natural killer (NK) cell activity of lymphocytes from healthy donors and from patients with the acquired immunodeficiency syndrome (AIDS). Two recombinant peptides (env-gag and Env 80-DHFR) and three chemically synthesized peptides (env 487-511, env 578-608 and env 647-659) were used. Normal lymphocytes precultured for 24 to 72 hrs. with either env-gag, env 487-511, or env 647-659 at 5 and 50 ng/ml concentrations which significantly stimulated lymphocyte proliferation, produced significant suppression of NK activities. Two control peptides, one derived from the E. coli vector used to clone the HIV env-gag fusion peptide and another, a non-HIV-1 viral antigen (rubeola virus) did not produce any observable effect on NK activity of normal lymphocytes demonstrating the specificity of the reaction. Env-gag peptide also inhibited the NK activities of Percoll-separated, NK-enriched large granular lymphocytes. In target binding assays, lymphocytes precultured with env-gag significantly suppressed the target binding capacity of effector cells and produced significantly lower levels of natural killer cytotoxic factor (NKCF). In kinetic studies, lymphocytes from normal donors preincubated with env-gag for 24 to 72 hrs. produced significant inhibition of their NK activity and an even greater inhibitory effect on NK activities was observed when lymphocytes from AIDS patients were preincubated with HIV peptides. Thus HIV-1 peptides, which we previously demonstrated could regulate B- and T-lymphocyte activities, are also capable of regulating the NK activities of lymphocytes from HIV-1-infected and normal individuals.
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PMID:Inhibition of natural killer cell activities from normal donors and AIDS patients by envelope peptides from human immunodeficiency virus type I. 944 29