ABSTRACT
Inactivation of mung bean glyceraldehyde-3-phosphate dehydrogenase (GPDH) with excess iodoacetate or N-ethylmaleimide exhibits pseudo-first order kinetics at pH 7.3 and 8.6 in the absence and presence of NAD+, suggesting that all the reactive SH groups (four per tetrameric GPDH molecule) have equivalent reactivity towards these reagents. This is similar to the D2-symmetry conformation proposed on the basis of thermal inactivation data [Malhotra and Srinivasan, Arch. Biochem. Biophys. 236, 775-781 (1985)]. With p-chloromercury benzoate (p-CMB), the inactivation of GPDH is very fast and its kinetics can be monitored at low reagent concentration only. Keeping a high molar p-CMB: enzyme ratio (= 47), the kinetics were found to be biphasic, with half of the activity being lost in a fast and the remaining in a slow phase, characteristic of C2-symmetry conformation and half site reactivity. The p-CMB inactivation could be largely reversed on the addition of excess cysteine. A comparison of these data with literature reports on this and other GPDHs reveals that all reagents having large non-polar moieties exhibit half site reactivity with this enzyme.
Subject(s)
Animals , Chloromercuribenzoates/pharmacology , Ethylmaleimide/pharmacology , Fabaceae/enzymology , Glyceraldehyde-3-Phosphate Dehydrogenases/antagonists & inhibitors , Kinetics , Plants/enzymology , Plants, Medicinal , Protein Conformation , Rabbits , Rats , Saccharomyces cerevisiae/enzymology , Sulfhydryl Reagents/pharmacology , Swine , p-Chloromercuribenzoic AcidABSTRACT
Parachloromercury benzoate (PCMB), a sulphydryl inactivator, caused a progressively increasing inhibition of tissue responses to acetylcholine, adrenaline, noradrenaline and serotonin in vitro. This inhibition was progressively and completely reversed by penicillamine, a sulphydryl activator. It is inferred that intact sulphydryl groups are essential for constancy of responses of excitable tissues to the neurotransmitters.
Subject(s)
Acetylcholine/pharmacology , Animals , Chloromercuribenzoates/pharmacology , Epinephrine/pharmacology , Muscle Contraction/drug effects , Norepinephrine/pharmacology , Penicillamine/pharmacology , Rabbits , Rats , Serotonin/pharmacology , Sulfhydryl Compounds/pharmacology , p-Chloromercuribenzoic AcidABSTRACT
From studies on the release of acid phosphatase and B-glucuronidase, it was observed that quercetin, a vitamin P like compound and ascorbic acid stablise while dehydroascorbate, acetoacetate and B-hydroxybutyrate labilise the leukocytic lysosomes in vitro. These effects were compared with chloroquine and progesterone, known stabiliser and destabiliser, respectively. The possible mode of lysosomal labilisation by ketone bodies and dehydroascorbate has been suggested.