Mutations that alter the charge of type I regulatory subunit and modify activation properties of cyclic AMP-dependent protein kinase from S49 mouse lymphoma cells

摘要: Abstract Mutations in regulatory (R) subunit of cAMP-dependent protein kinase were analyzed from cAMP-resistant mutants S49 mouse lymphoma cells by direct sequencing amplified regions mutant R cDNAs. Eight distinct single base-change lesions identified 24 independent that hemizygous for expression subunits with altered charge. CG----TA transitions predominated, but AT----GC and GC----TA transversions also observed. Four five spontaneous had identical C----T at CG causing substitution Trp Arg-334. Sites mutated isolates obtained after mutagenesis ethyl methanesulfonate or N-methyl-N'-nitro-N-nitrosoguanidine more varied. Six the (two binding site A four B) amino acid residues are highly conserved among cAMP-binding sites Escherichia coli catabolite activator protein. These mutations all either prevented strongly hindered cyclic nucleotides to site. One remaining (at Arg-242) nucleotide site; other Gly-170) only minimal effects on but, nevertheless, increased apparent constant activation. results discussed reference a model based crystal structure E.