Difference in kinetic properties between hexokinase type I isoenzymes from various rat tissues with reference to the effect of a thiol inhibitor (Short Communication)

Abstract

Hexokinase isoenzyme type I was purified from various rat tissues, and was subjected to kinetic studies in the presence or the absence of p-chloromercuribenzenesulphonate. The mode of the inhibition by the thiol inhibitor was different for the type I isoenzymes obtained from different tissues, suggesting that the type I isoenzymes from different tissues were not identical proteins.

Effect of ligands on the reactivity of essential sulfhydryls in brain hexokinase. Possible interaction between substrate binding sites

“…This phenomenon is explained by our observation that DTNB binds to the sites containing the sulfhydryls prior to the modification reaction. Our data showing competition between DTNB and the substrates to bind to the same site on the enzyme resemble the results obtained by Kamikashi et al (1974) with p-chloromercuribenzoate.…”

Discrimination of antigenic site and thiol-inhibitor-sensitive site of hexokinase isoenzymes

“…In other words, the extent of the inhibition was almost the same for all the type I isoenzymes examined. It should be pointed out that these type I isoenzymes were found in the previous studies to be different in their response to p-chloromercuribenzenesulphonate, both with respect to sensitivity and mode of inhibition (Murakami et al, 1973;Kamikashi et al, 1974;Ouchi et al, 1974). On the other hand, the activity of the hexokinase type II isoenzymes from skeletal muscle and spleen and that of the type III isoenzymes from spleen were not inhibited by the antiserum at all.…”

“…On the other hand, all of the type I isoenzymes were found to have the same antigenic site irrespective of the originating tissues and subcellular fractions, in spite of the dissimilarity of their response to the thiol inhibitor. The nature of the inhibition of hexokinase by the thiol inhibitor, especially of the competitive inhibition, has been discussed (Parry & Walker, 1967;Kamikashi et al, 1974) but has not been fully elucidated so far. If the competitive relationship between the substrate and inhibitor is, as discussed by Parry & Walker (1967), due to the effect of the substrate on the rate at which an enzyme thiol group undergoes an essentially irreversible reaction to form a covalently bonded complex with the inhibitor, such an inhibition may indicate the presence of, at least, a thiol group around the substrate-binding site.…”

“…The antiserum inhibited the activity of the mitochondrial hexokinase type I as well as that of the cytosolic type I enzyme prepared from rat brain, kidney and spleen. It (Schimke & Grossbard, 1968 (Murakami et al, 1973;Kamikashi et al, 1974). On the other hand, it is known that hexokinase in the brain is predominantly mitochondrial, though the enzyme is generally located in the cytosol in other tissues.…”