02035nas a2200229   4500008004100000022001400041245012600055210006900181260007500250300001200325490000700337520117600344653002701520653001601547653002001563653002701583653001401610653002301624100001801647700001401665856012601679       2013                        eng d  a1053-050900aMechanistic insights into the inhibition of endo-beta 1,4 xyloglucan hydrolase by a classical aspartic protease inhibitor0 aMechanistic insights into the inhibition of endobeta 14 xylogluc  a233 SPRING ST, NEW YORK, NY 10013 USAbSPRINGER/PLENUM PUBLISHERScMAR  a311-3210 v233 a<p>This is the first report of inactivation of xyloglucanase from Thermomonospora sp by pepstatin A, a specific inhibitor towards aspartic proteases. The steady state kinetics revealed a reversible, competitive, two-step inhibition mechanism with IC (50) and K (i) values of 3.5 +/- 0.5 mu M and 1.25 +/- 0.5 mu M respectively. The rate constants determined for the isomerization of EI to EI* and the dissociation of EI* were 14.5 +/- 1.5 x 10(-5) s(-1) and 2.85 +/- 1.2 x 10(-8) s(-1) respectively, whereas the overall inhibition constant K (i) (*) was 27 +/- 1 nM. The conformational changes induced upon inhibitor binding to xyloglucanase were monitored by fluorescence analysis and the rate constants derived were in agreement with the kinetic data. The abolished isoindole fluorescence of o-phthalaldehyde (OPTA)-labeled xyloglucanase and far UV analysis suggested that pepstatin binds to the active site of the enzyme. Our results revealed that the inactivation of xyloglucanase is due to the interference in the electronic microenvironment and disruption of the hydrogen-bonding network between the essential histidine and other residues involved in catalysis.</p>10a4 xyloglucan hydrolase10aEndo-beta 110aEnzyme kinetics10aInactivation mechanism10aPepstatin10aSlow-tight binding1 aMenon, Vishnu1 aRao, Mala  uhttps://library.ncl.res.in/content/mechanistic-insights-inhibition-endo-beta-14-xyloglucan-hydrolase-classical-aspartic-0