The protein p30, encoded at the gag-pro junction of mouse mammary tumor virus, is a dUTPase fused with a nucleocapsid protein.
A ribosomal frameshift at the gag-pro junction of mouse mammary tumor virus (MMTV) gives rise to the protein p30. The protein consists of two domains, the zinc-finger-containing nucleocapsid (NC) protein portion with 95 residues and a C-terminal extension comprising 154 residues. The C-terminal domain shows similarity in sequence with the enzyme dUTPase from other sources. In this paper, we demonstrate that p30 is a functional dUTPase. Overproduction of the NC protein in Escherichia coli, using the native frameshift sequence at the gag stop codon, caused a detectable expression of dUTPase ascribed to a low frequency of readthrough. By a 1-base insertion, eliminating the gag stop codon and fusing the gag and pro reading frames, a plasmid, pET-3d-NCDU, directing overexpression of p30, was constructed. The overproduced protein, purified by phosphocellulose chromatography, shows both zinc-binding and dUTPase activity. Analytical gel filtration and sequence homology to other dUTPases suggest a trimeric assembly of p30 subunits. MMTV thus possesses two different forms of the nucleocapsid protein, the ordinary NC protein and the p30, having the NC protein connected to a domain of dUTPase.
Bergman
A C
AC
Department of Biochemistry, University of Lund, Sweden.
Björnberg
O
O
Nord
J
J
Nyman
P O
PO
Rosengren
A M
AM
D000595:Amino Acid Sequence; D001483:Base Sequence; D002213:Capsid / Q000235:genetics / Q000378:metabolism*; D016368:Frameshift Mutation / Q000502:physiology; D015683:Gene Products, gag / Q000235:genetics*; D008324:Mammary Tumor Virus, Mouse / Q000737:chemistry* / Q000201:enzymology / Q000235:genetics; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011755:Pyrophosphatases / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D014758:Viral Core Proteins / Q000235:genetics / Q000378:metabolism*; D015032:Zinc / Q000378:metabolism
D015683:Gene Products, gag; D011993:Recombinant Fusion Proteins; D014758:Viral Core Proteins; D011755:Pyrophosphatases; C020020:dUTP pyrophosphatase; D015032:Zinc