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PMID: 2671992 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Isolation and expression of the Pneumocystis carinii thymidylate synthase gene.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 86 ·No. 17 ·1989-09-00 ·Pages 6503-7

Edman U, Edman JC, Lundgren B, Santi DV

Abstract

The thymidylate synthase (TS) gene from Pneumocystis carinii has been isolated from complementary and genomic DNA libraries and expressed in Escherichia coli. The coding sequence of TS is 891 nucleotides, encoding a 297-amino acid protein of Mr 34,269. The deduced amino acid sequence is similar to TS from other organisms and is most closely related to the enzyme from Saccharomyces cerevisiae with 65% identity. TS is found on a 330-kilobase-pair chromosome in P. carinii. While TS and dihydrofolate reductase reside on a single polypeptide chain in all protozoa studied to date, TS is not linked to dihydrofolate reductase in P. carinii. The TS gene shows the presence of four small intervening sequences, some of which interrupt the coding sequence in highly ordered structural regions of the protein. Heterologous expression of P. carinii TS in E. coli was accomplished by cloning the coding sequence into plasmid vectors under control of the lac and tac promoters. These constructs direct the synthesis of catalytically active enzyme to the extent of 2% of total soluble protein.

MeSH Terms
Amino Acid Sequence Animals Base Sequence DNA/genetics,isolation & purification Escherichia coli/genetics Gene Amplification Genes Molecular Sequence Data Oligonucleotide Probes Pneumocystis/enzymology,genetics Thymidylate Synthase/genetics
Chemicals
Oligonucleotide Probes DNA Thymidylate Synthase
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Edman U
Intercampus Program for Molecular Parasitology, University of California, San Francisco 94143.
Edman J C
Lundgren B
Santi D V
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Article Info
Journal
Proceedings of the National Academy of Sciences of the United States of America
Abbr.
Proc Natl Acad Sci U S A
ISSN
0027-8424
DOI
10.1073/pnas.86.17.6503
Published
1989-09-00
Pages
6503-7
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC297872
Subset
IM
Grants
NIAID NIH HHS · AI19358 · United States
NCI NIH HHS · CA19358 · United States
Databases
GENBANK
M25415, M25416
External Links
PubMed source DOI: 10.1073/pnas.86.17.6503 PMC Full Text View journal details
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