Nucleoside triphosphate-dependent DNA-binding properties of mos protein.

Abstract

We have previously shown that the mos gene product, p40f°, produced in Escherichia coli binds ATP and has ATPase activity. In the present study, we investigated the DNA-binding properties of p4011S and two mos deletion mutant proteins. Nitrocellulose blot protein-DNA binding assaysshowed that p40otm binds DNA in the presence of Mg2+-ATP and certain other nucleoside triphosphates. Ninety percent of the p40'°"-bound DNA is dissociated if the complex is washed in the presence of 1 M NaCl or in the absence of ATP. (4,5,11), the SV40 large T antigen shows both sequence-specific and nonspecific binding to DNA sequences (8)(9)(10)12). Recently, the DNA sequence-specific binding of SV40 large T antigen was shown to be abolished in an ATP-dependent manner by the allosteric effect of purine nucleoside triphosphates (13).However, in another case, the Tn3 transposase has been shown to bind specifically to the Tn3 inverted repeat in the presence of ATP (14). The viral mos (v-mos) gene of Moloney murine sarcoma virus (Mo-MSV) encodes a 37-kDa env-mos fusion protein (15,16). The mos proteins expressed by Mo-MSV strains tsllO and 124 have been shown to have serine/threonine autophosphorylation activity (17, 18). The v-mos product has been localized to the cytoplasm in acutely infected and transformed cells and is present at extremely low levels (19). The low level of mos product in transformed cells makes the characterization of its biochemical properties difficult. Therefore, we expressed the mos product and several mos deletion mutant proteins in Escherichia coli by using derivatives of high-level-expression vectors pJL6 and pANH-1 (20, 21). The derivative containing the complete mos gene (pA28) directed the synthesis of p40os (22), whereas those vectors lacking either N-terminal (pANHA28) or C-terminal (pA28A8) moieties expressed truncated versions of p4Omos, called pl9moS and p25m0s, respectively (21,22). The mos protein has strong homology with the ATP-binding domain of the cAMP-dependent bovine protein kinase and members of the src kinase family of oncogene products (23). We have shown that purified p40mOS expressed in E. coli binds ATP and has ATPase activity (22 (11,25). The purified mos and deletion mutant proteins were resolved in a NaDodSO4/12% polyacrylamide gel and were transferred electrophoretically onto nitrocellulose paper (26). The paper was incubated with 1% non-fat dry milk (27) in 0.5 M NaCl/50 mM Tris HCl, pH 8. 3560The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Mouse Mos protooncogene product is present and functions during oogenesis.

“…These studies have shown that it is a soluble cytoplasmic protein (21) with serine/threonine kinase activity (17,18). It also has been shown that the product of v-mos is an ATPase (19) and has ATP-dependent nucleic acid-binding activity (22). Both vmos and the mouse c-mos, designated Mos, have equivalent biological transforming activities in NIH 3T3 cell transfection assays (23).…”

The mos proto‐oncogene product: Its role in oocyte maturation, metaphase arrest, and neoplastic transformation

“…A number of studies have shown a perfect correlation between the protein kinase activity of both c-mos and v-mos and their biological function, either as tumor-causing proteins or as regulators of maturation-promoting factor (MPF) in oocytes and eggs [I , . Two other biochemical activities, namely an ATPase and a nucleic acid-binding activity, have been attributed to v-rnos [12,13]. However, the physiological relevance of these activities is currently unclear.…”

v-mos protein produced by in vitro translation has protein kinase activity

“…The v-mos protein, expressed in bacteria as a fusion protein, was found to have an ATP-dependent DNA-binding activity and an ATPase activity (22,23). This fusion protein, although expressed in large amounts in bacteria, was insoluble, requiring 8 M urea treatment for biochemical studies.…”