Phenotypic Expression inEscherichia coliand Nucleotide Sequence of Two Chinese Hamster Lung Cell cDNAs Encoding Different Dihydrofolate Reductases:

1984 ◽  
Vol 4 (5) ◽  
pp. 1001-1001
Keyword(s):  
Nucleotide Sequence ◽  
Phenotypic Expression ◽  
Chinese Hamster ◽  
Dihydrofolate Reductases

scholarly journals Phenotypic expression in Escherichia coli and nucleotide sequence of two Chinese hamster lung cell cDNAs encoding different dihydrofolate reductases.

1984 ◽  
Vol 4 (1) ◽  
pp. 38-48 ◽  
Author(s):  
P W Melera ◽  
J P Davide ◽  
C A Hession ◽  
K W Scotto
Keyword(s):  
Escherichia Coli ◽  
Molecular Weight ◽  
Nucleotide Sequence ◽  
Dihydrofolate Reductase ◽  
Phenotypic Expression ◽  
Chinese Hamster ◽  
Differential Sensitivity ◽  
Nucleotide Sequence Analysis ◽  
Cdna Clones ◽  
Weight Class

Nucleotide sequence analysis of two cDNA clones, one shown to direct the synthesis in Escherichia coli of the pI 6.7 form of the 20,000-molecular-weight class of Chinese hamster lung cell dihydrofolate reductase, and the other shown to direct the synthesis of the pI 6.5 form of the 21,000-molecular-weight class of the enzyme, has revealed the following: (i) the differences in physical and enzymatic properties displayed by these two proteins are due to two variations in their respective amino acid sequences with the conversion of Leu to Phe at position 22 probably responsible for the differential sensitivity of these two enzymes to methotrexate and methasquin; (ii) the multiple mRNAs responsible for the synthesis of each of these proteins differ in size due, at least in part, to a length heterogeneity at their 3' ends; (iii) these two proteins are encoded by different genes; and (iv) the sequence AAATATA appears to be a major polyadenylation signal in one Chinese hamster lung cell dihydrofolate reductase gene and a minor signal in another.

Phenotypic expression in Escherichia coli and nucleotide sequence of two Chinese hamster lung cell cDNAs encoding different dihydrofolate reductases

1984 ◽  
Vol 4 (1) ◽  
pp. 38-48
Author(s):  
P W Melera ◽  
J P Davide ◽  
C A Hession ◽  
K W Scotto
Keyword(s):  
Escherichia Coli ◽  
Molecular Weight ◽  
Nucleotide Sequence ◽  
Dihydrofolate Reductase ◽  
Phenotypic Expression ◽  
Chinese Hamster ◽  
Differential Sensitivity ◽  
Nucleotide Sequence Analysis ◽  
Cdna Clones ◽  
Weight Class

Nucleotide sequence analysis of two cDNA clones, one shown to direct the synthesis in Escherichia coli of the pI 6.7 form of the 20,000-molecular-weight class of Chinese hamster lung cell dihydrofolate reductase, and the other shown to direct the synthesis of the pI 6.5 form of the 21,000-molecular-weight class of the enzyme, has revealed the following: (i) the differences in physical and enzymatic properties displayed by these two proteins are due to two variations in their respective amino acid sequences with the conversion of Leu to Phe at position 22 probably responsible for the differential sensitivity of these two enzymes to methotrexate and methasquin; (ii) the multiple mRNAs responsible for the synthesis of each of these proteins differ in size due, at least in part, to a length heterogeneity at their 3' ends; (iii) these two proteins are encoded by different genes; and (iv) the sequence AAATATA appears to be a major polyadenylation signal in one Chinese hamster lung cell dihydrofolate reductase gene and a minor signal in another.

scholarly journals The Entire Nucleotide Sequence of Friend-Related and Paralysis-Inducing PVC-441 Murine Leukemia Virus (MuLV) and Its Comparison with Those of PVC-211 MuLV and Friend MuLV

1998 ◽  
Vol 72 (4) ◽  
pp. 3423-3426 ◽  
Author(s):  
Atsushi Tanaka ◽  
Kiyomasa Oka ◽  
Keiji Tanaka ◽  
Atsushi Jinno ◽  
Sandra K. Ruscetti ◽  
...  
Keyword(s):  
Nucleotide Sequence ◽  
Murine Leukemia Virus ◽  
Chinese Hamster Ovary Cells ◽  
Leukemia Virus ◽  
Chinese Hamster ◽  
Brain Capillary Endothelial Cell ◽  
Ovary Cells ◽  
The Difference ◽  
Entire Nucleotide Sequence ◽  
Murine Leukemia

ABSTRACT PVC-441 murine leukemia virus (MuLV) is a member of the PVC group of Friend MuLV (F-MuLV)-derived neuropathogenic retroviruses. In order to determine the molecular basis for the difference in neuropathogenicity between PVC-441 and the previously characterized PVC-211 MuLVs, the entire nucleotide sequence of PVC-441 MuLV was determined and compared with those of PVC-211 and F-MuLV. The results suggest that PVC-441 and PVC-211 MuLVs were formed as a result of random mutations of F-MuLV and developed differently. The distinct pathogenicities of PVC-441 and PVC-211 MuLVs were maintained in the viruses regenerated from their molecular clones, and the sequences responsible for the pathological differences observed can be localized to the env gene. The amino acid sequence of PVC-441 deduced from its nucleotide sequence revealed a number of differences from PVC-211, the most striking of which was a difference at position 129 of the SU proteins in the two viruses. Host range studies with a brain capillary endothelial cell line (RTEC-6) and Chinese hamster ovary cells (CHO-K1) revealed that PVC-441, like PVC-211, could infect these cells but its efficiency of infection was lower than that of PVC-211. These results may account for the difference in neuropathogenicity between PVC-441 and PVC-211.

scholarly journals Characterization of Dihydrofolate Reductase Genes from Trimethoprim-Susceptible and Trimethoprim-Resistant Strains ofEnterococcus faecalis

1999 ◽  
Vol 43 (1) ◽  
pp. 141-147 ◽  
Author(s):  
Teresa M. Coque ◽  
Kavindra V. Singh ◽  
George M. Weinstock ◽  
Barbara E. Murray
Keyword(s):  
Amino Acids ◽  
Nucleotide Sequence ◽  
E Coli ◽  
Integrative Vector ◽  
Dihydrofolate Reductases ◽  
Multiple Copies ◽  
Gram Negative Organisms ◽  
High Level

Enterococci are usually susceptible in vitro to trimethoprim; however, high-level resistance (HLR) (MICs, >1,024 μg/ml) has been reported. We studied Enterococcus faecalis DEL, for which the trimethoprim MIC was >1,024 μg/ml. No transfer of resistance was achieved by broth or filter matings. Two different genes that conferred trimethoprim resistance when they were cloned in Escherichia coli (MICs, 128 and >1,024 μg/ml) were studied. One gene that coded for a polypeptide of 165 amino acids (MIC, 128 μg/ml forE. coli) was identical to dfr homologs that we cloned from a trimethoprim-susceptible E. faecalis strain, and it is presumed to be the intrinsic E. faecalis dfr gene (which causes resistance in E. coli when cloned in multiple copies); this gene was designated dfrE. The nucleotide sequence 5′ to this dfr gene showed similarity to thymidylate synthetase genes, suggesting that the dfr andthy genes from E. faecalis are located in tandem. The E. faecalis gene that conferred HLR to trimethoprim in E. coli, designated dfrF, codes for a predicted polypeptide of 165 amino acids with 38 to 64% similarity with other dihydrofolate reductases from gram-positive and gram-negative organisms. The nucleotide sequence 5′ to dfrFdid not show similarity to the thy sequences. A DNA probe for dfrF hybridized under high-stringency conditions only to colony lysates of enterococci for which the trimethoprim MIC was >1,024 μg/ml; there was no hybridization to plasmid DNA from the strain of origin. To confirm that this gene causes trimethoprim resistance in enterococci, we cloned it into the integrative vector pAT113 and electroporated it into RH110 (E. faecalisOG1RF::Tn916ΔEm) (trimethoprim MIC, 0.5 μg/ml), which resulted in RH110 derivatives for which the trimethoprim MIC was >1,024 μg/ml. These results indicate thatdfrF is an acquired but probably chromosomally located gene which is responsible for in vitro HLR to trimethoprim in E. faecalis.

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