Bacteriophage T4 encodes an RNase H which removes RNA primers made by the T4 DNA replication system in vitro.
RNase H activity increases markedly after bacteriophage T4 infection of Escherichia coli MIC2003, an RNase H-deficient host. We have extensively purified the RNase H from these T4-infected cells and have shown that the RNase H activity copurifies with a 5' to 3' DNA exonuclease activity. The N-terminal sequence of a 35-kDa protein copurifying with the RNase H activity matches the terminus of the predicted product of an open reading frame (designated ORF A or 33.2) upstream of T4 gene 33, identified previously by Hahn and co-workers (Hahn, S., Kruse, U., and Rüger, W. (1986) Nucleic Acids Res. 14, 9311-9327). Plasmids containing ORF A under the control of the T7 promoter express RNase H and 5' to 3' DNA exonuclease activities as well as a protein that comigrates on sodium dodecyl sulfate-polyacrylamide gels with the 35-kDa protein present in the RNase H purified from T4-infected cells. T4 RNase H removes the pentamer RNA primers from DNA chains initiated by the T4 primase-helicase (gene products 61 and 41). Addition of T4 RNase H and T4 DNA ligase leads to extensive joining of discontinuous lagging strand fragments in the T4 DNA replication system in vitro.
Hollingsworth
H C
HC
Section on Nucleic Acid Biochemistry, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
Nossal
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NG
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D019915:DNA Primase; D004261:DNA Replication; D004279:DNA, Viral / Q000096:biosynthesis*; D004722:Endoribonucleases / Q000235:genetics / Q000378:metabolism*; D005090:Exodeoxyribonucleases / Q000378:metabolism; D005814:Genes, Viral; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009841:Oligonucleotides / Q000378:metabolism; D012313:RNA / Q000378:metabolism*; D012316:RNA Nucleotidyltransferases / Q000378:metabolism; D016914:Ribonuclease H; D013379:Substrate Specificity; D013604:T-Phages / Q000201:enzymology* / Q000235:genetics; D015678:Viral Structural Proteins / Q000235:genetics*; D014779:Virus Replication
D004279:DNA, Viral; D009841:Oligonucleotides; C034477:RNA primers; D015678:Viral Structural Proteins; D012313:RNA; D019915:DNA Primase; D012316:RNA Nucleotidyltransferases; D004722:Endoribonucleases; D005090:Exodeoxyribonucleases; D016914:Ribonuclease H