Characterization of a purified alpha-amylase inhibitor from white kidney beans (Phaseolus vulgaris)

A crude extract prepared from white kidney beans (Phaseolus vulgaris) showe d alpha-amylase inhibitory activity. Four fractions showed alpha-amylase in hibitory activity after the extract was subjected to reverse phase chromato graphy. The fraction with the highest activity was isolated and characteriz ed. It was found to be a glycoprotein with an N-glycosylation site whose de glycosylated molecular weight is 54,847 as determined by electrospray ioniz ation mass spectrometry (ESI-MS). Its C-terminal residues were serine, foll owed by alanine and tyrosine. Its binding constant was 2.81 mu M at 55 degr ees C. The carbohydrate moiety was not involved in binding as its removal d id not decrease the binding constant. Physiological amounts of kidney homog enate, plasma proteases, pronase or thermolysin readily hydrolysed the puri fied inhibitor; however, it was resistant to pepsin. Chloride ions were fou nd to be important for full activity while calcium increased the initial ra te of binding. Magnesium or sulfate ions did not have an effect on inhibiti on. Classical competitive inhibition of alpha-amylase was observed at pH 6. 9. (C) 1999 Canadian Institute of Food Science and Technology. Published by Elsevier Science Ltd. All rights reserved.