Regulation (alteration)of activity and conformationof sucrase by coaggregation with cellobiase in culture medium of termitomyces clypeatus

摘要

Extracellular sucrase(S)of Termitromyces clypeatus was aggregated with cellobiase (C) in culture filtrate and coaggregates of sucrase to cellobiase with different activity ratios(S/C)were obtained during purification.Specific activity of the enzyme decreased significantly after purification of sucrase free from cellobiase.Purified sucrase was characterized as a glycoprotein of molar mass around 55kDa as indicated by SDS-PAGE and HPGPLC.K_m and V_(max) of the purified enzyme were determined as 34.48mM and 13.3U/mg,respectively,at optimum temperature (45degC) and pH(5.0).Substrate affinity and reaction velocity of the purified enzyme,free from cellobiase,was lowered by approximately 3,5 and 55 yimes,respectively,than that of the enzyme obtained from culture filtrate.The instant ragain of sucrase activity up to the extent of 41% was obtained on in vitro addition of cellobiase(free from sucrase)to the enzyme in incubation mixtrue.Conformationof the enzyme free from cellobiase appeared to be significantly different from that of the coaggregate,as analyzed by circular dichroic and light scattering spectroscopy.It was concluded that activity and conformation of sucrase is regulated (altered)by heteroaggregation with cellobiase in the fungus.