Kinetics and mechanism of the hydrolysis of sodium carboxymethylcellulose (Na-CMC) by a cellulase complex

TheSomogyi-Nelson colorimetric method is applied in a new manner more suitable for evaluating the kinetics of the enzyme hydrolysis of sodium carboxymethylcellulose (Na-CMC) catalyzed by the cellulase complex. By means of selective inhibition of a chosen enzyme from the cellulase complex it became possible to trace the effect of the other enzymes included in its composition.

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Kinetik und Mechanismus der Hydrolyse von Natriumcarboxymethylcellulose (Na-CMC) durch einen Cellulase-Komplex

Zusammenfassung Die kolorimetrische Methode nachSomogyi undNelson wird nach einem neuen Verfahren zur Verfolgung der Kinetik der hydrolytischen Spaltung von Natriumcarboxymethylcellulose (Na-CMC), katalysiert durch den Cellulase-Komplex, angewandt. Durch selektive Inhibierung eines bestimmten Enzyms des Cellulase-Komplexes kann man die Wirkung der anderen zu seiner gesamten Zusammensetzung gehörenden Enzyme verfolgen.

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Symbols Used E enzyme (E3735x70768/xxlarge8242.gif" alt="prime" align="BASELINE" BORDER="0">—cellulase;E3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0">—exo-cellobiohydrolase;E3735x70768/xxlarge8244.gif" alt="tprime" align="MIDDLE" BORDER="0">—3735x70768/xxlarge946.gif" alt="beta" align="MIDDLE" BORDER="0">-glucosidase) - [E] _w weight concentration of enzymeE - S substrate (Na-CMC—sodium carboxymethylcellulose) - [S]₀ weight concentration of substrateS - I inhibitor (I3735x70768/xxlarge8242.gif" alt="prime" align="BASELINE" BORDER="0">—lactose;I3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0">—calcium chloride;I3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0">—condurrite-B-epoxide) - P product (P3735x70768/xxlarge8242.gif" alt="prime" align="BASELINE" BORDER="0">—oligosaccharides;P3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0">—cellobiose;P3735x70768/xxlarge8244.gif" alt="tprime" align="MIDDLE" BORDER="0">—D-glucose) - P _{3735x70768/xxlarge8733.gif" alt="prop" align="MIDDLE" BORDER="0">} end product (K _{3735x70768/xxlarge8733.gif" alt="prop" align="MIDDLE" BORDER="0">} ^{3735x70768/xxlarge8242.gif" alt="prime" align="BASELINE" BORDER="0">} , K _{3735x70768/xxlarge8733.gif" alt="prop" align="MIDDLE" BORDER="0">} ^{3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0">} , K _{3735x70768/xxlarge8733.gif" alt="prop" align="MIDDLE" BORDER="0">} ^{3735x70768/xxlarge8244.gif" alt="tprime" align="MIDDLE" BORDER="0">} ) - DP degree of polymerization - DS degree of substitution - ES enzyme-substrate complex (E3735x70768/xxlarge8242.gif" alt="prime" align="BASELINE" BORDER="0"> S, E3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0"> S, E3735x70768/xxlarge8244.gif" alt="tprime" align="MIDDLE" BORDER="0"> S) - EP enzyme-product complex (E3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0"> P3735x70768/xxlarge8242.gif" alt="prime" align="BASELINE" BORDER="0">, E3735x70768/xxlarge8244.gif" alt="tprime" align="MIDDLE" BORDER="0"> P3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0">) - EI enzyme-inhibitor complex (E3735x70768/xxlarge8242.gif" alt="prime" align="BASELINE" BORDER="0"> I3735x70768/xxlarge8242.gif" alt="prime" align="BASELINE" BORDER="0">, E3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0"> I3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0">, E3735x70768/xxlarge8244.gif" alt="tprime" align="MIDDLE" BORDER="0"> I3735x70768/xxlarge8244.gif" alt="tprime" align="MIDDLE" BORDER="0">) - M _s molecular mass of substrateS - K _s substrate constant (K _s ^{3735x70768/xxlarge8242.gif" alt="prime" align="BASELINE" BORDER="0">} , K _s ^{3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0">} , K _s ^{3735x70768/xxlarge8244.gif" alt="tprime" align="MIDDLE" BORDER="0">} ) - K _I inhibitor constant (K _I ^{3735x70768/xxlarge8242.gif" alt="prime" align="BASELINE" BORDER="0">} , K _I ^{3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0">} , K _I ^{3735x70768/xxlarge8244.gif" alt="tprime" align="MIDDLE" BORDER="0">} ) - K _m Michaelis-Menten constant - k ₊₁,k ₊₂ (k ₊₂ ^{3735x70768/xxlarge8242.gif" alt="prime" align="BASELINE" BORDER="0">} ,k ₊₂ ^{3735x70768/xxlarge8243.gif" alt="Prime" align="MIDDLE" BORDER="0">} ,k ₊₂ ^{3735x70768/xxlarge8244.gif" alt="tprime" align="MIDDLE" BORDER="0">} ) forward rate constants - k _–1 reverse rate constant - 3735x70768/xxlarge957.gif" alt="ngr" align="BASELINE" BORDER="0"> ₀ initial rate of reaction - V maximal reaction rate - 3735x70768/xxlarge916.gif" alt="Delta" align="BASELINE" BORDER="0">A change in absorbance - 3735x70768/xxlarge949.gif" alt="epsi" align="BASELINE" BORDER="0"> molar absorption coefficient - 3735x70768/xxlarge955.gif" alt="lambda" align="BASELINE" BORDER="0"> wavelength Herrn Prof. Dr.Hans Tuppy zum 60. Geburtstag herzlichst gewidmet.