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Tertiary structure‐dependence of misfolding substitutions in loops of the maltose‐binding protein
Author(s) -
Raffy Sébastien,
Sassoon Nathalie,
Hofnung Maurice,
Betton JeanMichel
Publication year - 1998
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.5560071010
Subject(s) - folding (dsp implementation) , maltose binding protein , periplasmic space , protein folding , chemistry , context (archaeology) , biophysics , loop (graph theory) , amino acid , protein tertiary structure , crystallography , escherichia coli , biochemistry , biology , gene , paleontology , mathematics , engineering , combinatorics , electrical engineering , fusion protein , recombinant dna
We previously identified and characterized amino acid substitutions in a loop connecting helix I to strand B, the αI/βB loop, of the N‐domain that are critical for in vivo folding of the maltose‐binding protein (MalE31). The tertiary context‐dependence of this mutation in MalE folding was assessed by probing the tolerance of an equivalent αβ loop of the C‐domain to the same amino acid substitutions (MaIE219). Moving the loop mutation from the N‐ to the C‐domain eliminated the in vivo misfolding step that led to the formation of inclusion bodies. In vitro, both loop variants exhibited an important decrease of stability, but their intrinsic tendency to aggregate was well correlated with their periplasmic fates in Escherichia coli . Furthermore, the noncoincidence of the unfolding and refolding transition curves and increase of light scattering during the refolding of MalE31 indicate that a competing off‐pathway reaction could occurs on the folding pathway of this variant. These results strongly support the notion that the formation of super‐secondary structures of the N‐domain is a rate‐limiting step in the folding pathway of MalE.