Molecular properties of global suppressors of temperature-sensitive folding mutations in P22 tailspike endorhamnosidase.
Two global suppressors (Val-331 greater than Ala and Ala-334 greater than Val) have been identified for temperature-sensitive folding (tsf) mutations in gene 9 of bacteriophage P22 (Mitraki, A., Fane, B., Haase-Pettingell, C., Sturtevant, J., and King, J. (1991) Science 253, 54-58). We have introduced 19 different single amino acid substitutions at the two global suppressor sites independently and examined the effects on the tailspike formation in Escherichia coli. Folding and maturation patterns of the various substitutions at the two global suppressor sites in the wild-type background suggest that Val-331 is located on the protein surface and Ala-334 is in the hydrophobic region. In combination with a tsf mutation, tsfH304 (Gly-244 greater than Arg), only Gly at 331 and Ile at 334, the substitutions that have similar side chain properties to the original suppressor sequences, were active as tsf suppressors. The newly identified suppressors of tsfH304 could also alleviate the tsf defect of three other mutations. The mutant carrying both Val-331 greater than Ala and Ala-334 greater than Val substitutions was also a global suppressor and was more active in suppressing the tsf defect than mutants carrying only one substitution. The suppressors may act by increasing the stability of an intermediate in the productive pathway of folding and maturation of the mutant polypeptides.
Lee
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Genetic Engineering Research Institute, Korea Institute of Science and Technology, Daeduck Science Town, Daejeon.
Koh
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Yu
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D000596:Amino Acids / Q000737:chemistry; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D013489:Suppression, Genetic; D013696:Temperature; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017376:Viral Tail Proteins
D000596:Amino Acids; D014764:Viral Proteins; D017376:Viral Tail Proteins; D006026:Glycoside Hydrolases; C041312:endorhamnosidase