Open AccessDirect Participation of dCMP Hydroxymethylase in Synthesis of Bacteriophage T4 DNA
Author(s) -
Merle G. Wovcha,
Paul K. Tomich,
C S Chiu,
Gordon R. Greenberg
Publication year - 1973
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.70.8.2196
Subject(s) - dna synthesis , biology , dna , microbiology and biotechnology , bacteriophage , mutant , dna replication , deoxyribonucleotide , gene , biochemistry , dnag , oligonucleotide , escherichia coli , eukaryotic dna replication
In order to retain in anin situ system the control mechanisms involved in synthesis of bacteriophage T4 DNA, infected cells were made permeable to nucleotides by plasmolysis with concentrated sucrose. Such preparations use exogenous deoxyribonucleotides to synthesize T4 phage DNA. As has been observed within vivo studies, DNA synthesis was drastically reduced in plasmolyzed preparations from cells infected byamber mutants of genes 1, 32, 41, 42, 43, 44, or 45. Added 5-hydroxymethyl dCTP did not bypass either a mutant of gene 42 (dCMP hydroxymethylase) or of gene 1 (phage-induced deoxyribonucleotide kinase). In a phage system lacking deoxycytidine triphosphatase (gene 56) and the gene-46 product, and therefore incorporating dCTP into DNA, dCTP incorporation did not require dCMP hydroxymethylase, in keeping within vivo results. With a tripleamber mutant of genes 1, 46, and 56 only slight incorporation of dCTP occurred. By contrast, in experiments performedin vivo the synthesis of cytosine-containing DNA was unaffected by anamber mutation in gene 1.These studies provide evidence that dCMP hydroxymethylase, in addition to its known catalytic function, has a second, more direct, role in phage T4 DNA synthesis, apparently in recognition of hydroxymethyl dCTP. The role of the phage-induced deoxyribonucleotide kinase in T4 DNA synthesis in the plasmolyzed system remains unresolved.