Asparagine 26, glutamic acid 31, valine 45, and tyrosine 64 of Ras proteins are required for their oncogenicity.

M S Nur-E-Kamal,A Sizeland,G D'Abaco,H Maruta

doi:10.1016/s0021-9258(18)45958-3

M S Nur-E-Kamal, A Sizeland + Show 2 more

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https://doi.org/10.1016/s0021-9258(18)45958-3

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Abstract

Ras and Rap1 proteins are related GTP-dependent signal transducers which require Gly-12, the effector domain (residues 32-40), and Ala-59 for stimulation of their GTPase activities by GAP1 and GAP3, respectively. The replacement of Gly-12 by Val or Ala-59 by Thr potentiates the Ras oncogenicity and Rap1A antioncogenicity. However, the mutations in the effector domain, in particular the replacement of Thr-35 by Ala, abolish both Ras oncogenicity and Rap1A antioncogenicity, indicating that the effector domain is involved in interactions of these signal transducers with their targets as well as the GAPs. In this paper, we demonstrate that (i) replacement of Tyr-64 of the Ha-Ras protein or Phe-64 of the Rap1A protein by Glu or other non-hydrophobic amino acids reduces their intrinsic GTPase activities and abolishes their stimulation by GAP1 or GAP3, respectively, (ii) replacement of Tyr-64 by Gly and other non-hydrophobic amino acids results in complete loss of the oncogenicity of the v-Ha-Ras protein, indicating that the hydrophobic residue 64, in addition to the known effector domain, is essential for the Ras protein to interact with its target as well as GAP1. In addition we have found that Asn-26, Glu-31, and Val-45 of the v-Ha-Ras protein are required for its oncogenicity. Replacement of the Ras residues at either positions 26, 31, or 45 by the corresponding Rap1A residues abolishes the Ras oncogenicity.

Highlights

We demonstrate that (i) replacement of Tyr-64 of the Ha-Ras protein or Phe-64 of the RaplA protein by Glu or other non-hydrophobic amino acids reduces their interactionswithGAPland GAP3, respectively
Since Gln-61 is not required for the Ras-dependent signal transduction (13),replacement of Gln[61] by Leu or other amino acids significantly potentiates the Ras oncogenicity (1).replacement of Thr-61 by Gln-61 in the RaplA protein significantlyreduces its antioncogenicity (6),althoughthis does not affect eitherthe intrinsic GTPase activity or its stimulation by GAP3 (12)
Intrinsic GTPase activities and abolishes their stimu- replacementof Gln-63 by Glu significantly relation byGAPl or GAP3, respectively, (ii)replacement duces theRaplGTPaseactivity (12) andpotentiatesthe of Tyr-64 by Gly and other non-hydrophobic amino acids resultsin complete loss of the oncogenicity of the v-Ha-Ras protein, indicating that the hydrophobic residue 64, in addition to the known effector domain, is essential for the Ras protein to interact with its target as well as GAP1

Summary

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Interest- Ala-66 by either Gly, Trp, Glu, or Arg and Glu-62 by Ala had ingly, the Phe-64 mutant of the Ha-Ras GTPase was not no significant effect on the enzymatic property of the Ras significantly activated by GAP3, and the Tyr-64 mutant of protein (Table 11). These observations confirm that Tyr or the Rapl GTPase was notactivated by GAPlC (data not Phe apt osition 64 plays aunique role in thesignal attenuation shown). None of the position 64 mutants derived from c-Ha-Ras gene allows the

Ras OncogenicDeterminants

Colonies formed in soft agar