Inhibition of cellobiohydrolase I from Trichoderma reesei by palladium.
Cellulase from Trichoderma reesei is a multienzyme mixture that hydrolyzes cellulose to glucose. Two enzymes in this mixture, cellobiohydrolase (CBH) and endoglucanase (EG), possess a common structure comprising a distinct cellulose-binding domain (CBD) and catalytic domain. Inhibition of the catalytic domain of cellulases without affecting their CBD function might be useful for structure/function studies of these enzymes. Complexes of the platinum group metals were tested for their ability to inhibit the major cellulase enzyme from T. reesei, cellobiohydrolase I (CBH I). Only palladium complexes inhibited CBH I, inhibition being dependent upon the molar ratio of palladium to CBH I with 1 microM CBH I retaining only 10% of its activity in the presence of 100 microM ammonium hexachloropalladate(IV) and after the incorporation of 28 mol Pd/mol CBH I. Inhibition was irreversible and could be completely prevented by including histidine, cysteine, and cystine in the assay mixture. Although the primary mechanism of inhibition of CBH I by palladium remains to be elucidated, it could involve the binding of palladium to sulfur or cystine residues resulting in their degradation. This is based on the findings that (i) palladium-inhibited CBH I was less thermally stable than native CBH I; (ii) CBH I, chemically modified by the attachment of pentaammine ruthenium(III) to the imidazole-N of either H206 or H228, showed greater sensitivity to inhibition by palladium compared to native CBH I; and (iii) ammonium hexachloropalladate cleaved 5,5'-dithiobis(2-nitrobenzoic acid)--Ellman's reagent. Binding of CBH I to crystalline cotton linters was not affected by palladium.
Lassig
J P
JP
Chemical Technology Division, Oak Ridge National Laboratory, Tennessee 37831-6194, USA.
Shultz
M D
MD
Gooch
M G
MG
Evans
B R
BR
Woodward
J
J
D000596:Amino Acids / Q000494:pharmacology; D001665:Binding Sites; D002482:Cellulose / Q000378:metabolism; D043366:Cellulose 1,4-beta-Cellobiosidase; D002942:Circular Dichroism; D004220:Disulfides / Q000737:chemistry; D006026:Glycoside Hydrolases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D006358:Hot Temperature; D007700:Kinetics; D008670:Metals / Q000494:pharmacology; D010165:Palladium / Q000494:pharmacology*; D010984:Platinum / Q000494:pharmacology; D011489:Protein Denaturation; D012428:Ruthenium / Q000494:pharmacology; D013053:Spectrophotometry; D014242:Trichoderma / Q000201:enzymology*
D000596:Amino Acids; D004220:Disulfides; D008670:Metals; D010984:Platinum; D010165:Palladium; D012428:Ruthenium; D002482:Cellulose; D006026:Glycoside Hydrolases; D043366:Cellulose 1,4-beta-Cellobiosidase