Microassay for rapid screening of alpha-amylase activity.
A microassay was developed for measuring the activity of alpha-amylases in the nanogram enzyme concentration range, based on the use of dye-labeled cross-linked starch as the substrate, and the release of soluble colored fragments formed in enzyme hydrolysis. Reaction conditions were optimized to generate a linear correlation between the increase in absorbance and a reaction time of 0-10 min, as well as enzyme concentrations in the range of 0-50 ng. A standard curve for the conversion of absorbance to enzyme activity units was constructed. The protocol developed was applied to monitoring the production of ultralow concentrations of recombinant barley alpha-amylase in yeast cells.
Wong
D W
DW
Western Regional Research Center, Agricultural Research Service, U.S. Department of Agriculture, 800 Buchanan Street, Albany, California 94710, USA. dwsw@pw.usda.gov
Batt
S B
SB
Robertson
G H
GH
D004396:Coloring Agents; D007527:Isoenzymes / Q000737:chemistry; D008832:Microchemistry; D011994:Recombinant Proteins / Q000737:chemistry; D013213:Starch; D000516:alpha-Amylases / Q000737:chemistry*
D004396:Coloring Agents; D007527:Isoenzymes; D011994:Recombinant Proteins; D013213:Starch; D000516:alpha-Amylases