Properties of the nuclear P1 protein, a mammalian homologue of the yeast Mcm3 replication protein.
Polyclonal antibodies were raised against a multiprotein 'holoenzyme' form of calf thymus DNA polymerase alpha-primase and used to probe a human cDNA-protein expression library constructed in the lambda gt11 vector. The probe identified a series of cDNA clones derived from a 3.2 kb mRNA which encodes a novel 105 kDa polypeptide, the P1 protein. In intact cells, the P1 protein was specifically associated with the nucleus, and in cell extracts, it was associated with complex forms of DNA polymerase alpha-primase. The synthesis of human P1-specific mRNA was stimulated upon addition of fresh serum to growth-arrested cells, and RNA blot analyses with the human P1-cDNA probe indicated that P1 is encoded by a strictly conserved mammalian gene. The amino acid sequence deduced from a 240-codon open reading frame resident in the largest human P1-cDNA (0.84 kb) displayed greater than 96% identity with that deduced from the equivalent segment of a 795-codon open reading frame of a larger mouse P1-cDNA (2.8 kb). Throughout its length, the primary structure of mammalian P1 displayed strong homology with that of Mcm3, a 125 kDa yeast protein thought to be involved in the initiation of DNA replication (Gibson et al. 1990. Mol. Cell. Biol. 10: 5707-5720). The P1-Mcm3 homology, the strong conservation of P1 among mammals, its nuclear localization, and its association with the replication-specific DNA polymerase alpha strongly suggest an important role of the P1 protein in the replication of mammalian DNA.
Thömmes
P
P
Division of Biology, Universität Konstanz, FRG.
Fett
R
R
Schray
B
B
Burkhart
R
R
Barnes
M
M
Kennedy
C
C
Brown
N C
NC
Knippers
R
R
D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D018797:Cell Cycle Proteins; D002460:Cell Line; D003001:Cloning, Molecular; D019915:DNA Primase; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D051379:Mice; D064126:Minichromosome Maintenance Complex Component 3; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012316:RNA Nucleotidyltransferases / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid; D014157:Transcription Factors
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C066470:MCM3 protein, human; C513269:Mcm3 protein, mouse; D009687:Nuclear Proteins; D014157:Transcription Factors; D019915:DNA Primase; D012316:RNA Nucleotidyltransferases; D064126:Minichromosome Maintenance Complex Component 3