Biomedical subjects
M Innis
Publications and source records attributed to M Innis.
Biochemical and biological properties of the human N-ras p21 protein.
We characterized the normal (Gly-12) and two mutant (Asp-12 and Val-12) forms of human N-ras proteins produced by Escherichia coli. No significant differences were found between normal and mutant p21 proteins in their affinities for GTP or GDP. Examination of GTPase activities revealed significant differences between the mutant p21s: the Val-12 mutant retained 12% of wild-type GTPase activity, whereas the Asp-12 mutant retained 43%. Both mutant proteins, however, were equally potent in causing morphological transformation and increased cell motility after their microinjection into quiescent NIH 3T3 cells. This lack of correlation between transforming potency and GTPase activity or guanine nucleotide binding suggests that position 12 mutations affect other aspects of p21 function.
Genetic and biochemical analysis of ras p21 structure.
We tested aspects of our model of the ras p21 structure using generic, biochemical, and immunologic approaches. First, we made a monoclonal antibody against a p21 region that is highly conserved and likely to be critical to p21 function. The antibody blocks p21 function in various cell systems. Its binding to p21 is completely blocked by guanine nucleotides, even though the region of p21 to which it binds does not seem to be part of the guanine nucleotide-binding site. We propose that the conformation of this critical region is modulated by nucleotide binding. Another interesting region of p21 includes amino acids 116 and 119, which seem to confer, in part, the specificity of p21 for guanine nucleotides. We made a series of mutants in this region and tested their ability to bind GTP, and such related purine nucleotides as XTP and diaminopurine nucleoside triphosphate. We were able to refine our model for guanine nucleotide interaction with p21 and to create mutant proteins with altered specificity for purine nucleotides. Finally, we tested rates of autophosphorylation of six position 12 mutants and conclude that amino acid 12 affects the positioning of bound nucleotides relative to sequences around amino acid 59.
Detection of activated Mr 21,000 protein, the product of ras oncogenes, using antibodies with specificity for amino acid 12.
Antisera raised to a set of chemically synthesized peptides spanning position 12 of ras Mr 21,000 protein (p21) (residues 5 to 17) were able to distinguish between different forms of p21 according to the amino acid at the twelfth codon. The peptide immunogens differed in one amino acid corresponding to position 12 of the protein; the substitutions were valine, serine, arginine, aspartate, alanine, or cysteine at this position. Normal p21 contains glycine at position 12; the other amino acid substitutions are those which would result from a single base change in codon 12 and may therefore be the activating mutations most likely to occur in human tumors. The peptide antisera were evaluated by the Western immunoblot procedure for reactivity with v-ki-ras p21 expressed in Escherichia coli containing the corresponding position 12 mutations. Five of the antisera reacted with p21, and of these, anti-serine, -valine, -arginine, and -aspartate peptide antibodies were specific for their cognate protein. Similar analysis using mammalian cells as sources of position 12 variant forms of p21 demonstrated the ability of these antisera to distinguish among their oncogenic forms of p21 differing by single amino acid substitutions.
Factors likely to affect the development of multiple sclerosis in patients presenting with optic neuritis in a tropical and subtropical area.
The relationship between optic neuritis (ON) and multiple sclerosis (MS) in a subtropical climate is examined. 105 cases of ON were followed for varying periods over 26 years. The factors studied included place of birth, sex, age at onset, presence of recurrent ON, the interval to the onset of MS, and HLA antigen typing. 55% of cases developed evidence of definite MS. The majority of cases who went on to develop MS had their onset of ON between 21 and 40 years of age. The subtropical climate did not cause any significant variation in incidence. However those case who had HLA Bw4 appear to be protected from developing MS.