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THE MOLECULAR WEIGHT OF α‐AMYLASE INHIBITOR FROM WHITE BEAN cv 858B ( PHASEOLUS VULGARIS L.) IS 56 kDa, NOT 20 kDa
Author(s) -
LEE SHIHCHIEH,
WHITAKER JOHN R.
Publication year - 2000
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.1745-4514.2000.tb00043.x
Subject(s) - size exclusion chromatography , sephadex , phaseolus , chemistry , polyacrylamide gel electrophoresis , chromatography , gel electrophoresis , molecular mass , sodium dodecyl sulfate , cultivar , biochemistry , enzyme , biology , botany
The molecular weights (M r s) of α‐amylase inhibitors (αAIs)fiom 18 (Ah) bean cultivars estimated by Superose 12 gelfiltration chromatography were 22–62% smaller than those determined by Sephadex G‐100 gel filtration and by polyacrylamide gel electrophoresis (PAGE) methods. αAI‐4 from WKB cultivar 858B was purified and the Mr was shown to be 51.0 kDa based on Sephadex G‐100 gel filtration chromatography and by PAGE. A M r for aAI‐4 of 56.714 kDa was determined by laser‐assisted time‐of‐flight mass spectrometry and appears to be the true M r of the mature glycosylated active aAI‐4. The results show that Superose 12 gelfiltration chromatography is not usefulfor M r determination of some proteins. Sodium dodecyl sulfate electrophoresis (SDS‐PAGE) showed that the 56.7 kDa aAI‐4 molecule dissociated into 45.0, 33.6,15.2 and 12.4 kDa submolecules, with only the two small subunits, a and β, present at high SDS concentration. This provides evidence that the aAI‐4 molecules composition is α 2 β 2 .