title: Formation of aggregates from a thermolabile in vivo folding intermediate in P22 tailspike maturation. A model for inclusion body formation.
abstract: The in vivo accumulation of polypeptide chains in the form of aggregated non-native states is a problem in many applications of biotechnology. In the maturation pathway of the thermostable P22 tailspike endorhamnosidase, the folding and chain association intermediates can be distinguished from the native tailspikes in crude extracts of phage-infected Salmonella cells. Temperature-sensitive folding mutations, at many sites in the chain, destabilize these conformational intermediates preventing the formation of native tailspikes at restrictive temperatures (Goldenberg, D. P., Smith, D. H., and King, J. (1983) Proc. Natl. Acad. Sci. U. S. A. 80, 7060-7064). We report here that both wild type and mutant tailspike polypeptide chains which fail to reach the native state accumulate in an aggregated state. These off-pathway aggregates form from a thermolabile intermediate in the productive folding pathway. These aggregation reactions are suppressed by lowering the temperature of maturation. Similar off-pathway steps from folding intermediates may account for the non-native aggregates often found in the expression of cloned genes in heterologous hosts.
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
authors.forename: C A
authors.initials: CA
authors.lastname: Haase-Pettingell
authors.forename: J
authors.initials: J
authors.lastname: King
mesh_terms: D007700:Kinetics; D046911:Macromolecular Substances; D009154:Mutation; D012483:Salmonella Phages / Q000235:genetics / Q000378:metabolism*; D012486:Salmonella typhimurium / Q000235:genetics / Q000378:metabolism*; D013696:Temperature; D014357:Trypsin; D014764:Viral Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017376:Viral Tail Proteins
chemical_list: D046911:Macromolecular Substances; D014764:Viral Proteins; D017376:Viral Tail Proteins; D014357:Trypsin