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PERTURBATION OF THE STRUCTURE OF CLOSTRIDIUM PERFRINGENS ENTEROTOXIN BY SODIUM DODECYL SULFATE, GUANIDINE HYDROCHLORIDE, pH AND TEMPERATURE
Author(s) -
GRANUM PER EINAR,
WHITAKER JOHN R.
Publication year - 1980
Publication title -
journal of food biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.507
H-Index - 47
eISSN - 1745-4514
pISSN - 0145-8884
DOI - 10.1111/j.1745-4514.1980.tb00782.x
Subject(s) - chemistry , guanidine , sodium dodecyl sulfate , dtnb , hydrochloride , sodium , tryptophan , clostridium perfringens , nuclear chemistry , amino acid , inorganic chemistry , chromatography , biochemistry , organic chemistry , enzyme , bacteria , glutathione , biology , genetics
The effects of sodium dodecyl sulfate, guanidine · HCl, heat and pH on the tertiary structure of Clostridium perfringens type A enterotoxin were determined by UV difference spectroscopy and by accessibility of the amino groups to 2, 4, 6‐trinitrobenzene sulfonic acid (TNBS) and the one sulfhydryl group to 5, 5′‐dithiobis (2‐nitrobenzoic acid) (DTNB). Biological stability of enterotoxin at several pH's at 55°C in the presence and absence of borate and dipicolinate ions was determined by the guinea pig skin test. Sodium dodecyl sulfate and heat did not unfold the molecule while treatment with >4.5 M guanidine · HCl at ≥25°C completely unfolded the molecule based on UV difference spectral changes and the accessibility of the amino groups to reaction with TNBS. Sodium dodecyl sulfate, at 0.2%, made the sulfhydryl group accessible to DTNB but did not affect the accessibility of the amino groups (7.89 ± 0.15 groups in the native molecule). Guanidine · HCl, at 6.5 M for 30 min and ≥25°C, made all 17 e‐amino groups and the one α‐amino group accessible to TNBS. At pH's above 8 spectral differences, in relation to the spectrum at pH 6.8, were associated primarily with ionization of the tyrosine residues while at pH's below 5.5 the spectral changes were probably a result of protonation of carboxyl groups in the vicinity of tyrosine and tryptophan residues, and of aggregation.