Inhibition of phosphomannose isomerase by mercury ions.
Mercury ions can inhibit Candida albicans phosphomannose isomerase (PMI) by two different processes at sub-micromolar concentrations. Kinetic studies show that mercury ions are in rapid equilibrium with the enzyme and cause a clear partial noncompetitive inhibition when mannose 6-phosphate is used as the substrate. The inhibition constants at 37 degrees C in 50 mM Hepes buffer, pH 8.0, are 35 and 57 nM for Kii and Kis, respectively. In addition to this inhibition at rapid equilibrium, mercury ions also inactivate C. albicans PMI by a much slower process, involving an irreversible mechanism. This is shown to be a two-step process, proceeding via an intermediate complex with a dissociation constant of 5.6 microM, with a maximum rate of inactivation of 0.15 min-1. The rate of irreversible inactivation can be slowed by the addition of the substrate, mannose 6-phosphate. Incubation of the enzyme with [203Hg]Cl2 causes the formation of a stable adduct with one atom of mercury incorporated into each enzyme molecule during the inactivation. Since cysteine-150 is the only iodoacetate-modifiable cysteine in the protein, we propose that this is where the mercury ion reacts during the irreversible inactivation process. In the Escherichia coli enzyme this cysteine is replaced by an asparagine, and the enzyme cannot be irreversibly inactivated by mercury ions.(ABSTRACT TRUNCATED AT 250 WORDS)
Glaxo Institute for Molecular Biology, Geneva, Switzerland.
T N
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Wells
M A
MA
Payton
A E
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Proudfoot
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002176:Candida albicans / Q000201:enzymology; D003545:Cysteine / Q000737:chemistry; D004926:Escherichia coli / Q000201:enzymology; D006801:Humans; D007700:Kinetics; D008359:Mannose-6-Phosphate Isomerase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D008627:Mercuric Chloride / Q000494:pharmacology; D017669:Mercury Compounds / Q000378:metabolism / Q000494:pharmacology*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D013045:Species Specificity; D013552:Swine
D017669:Mercury Compounds; D011994:Recombinant Proteins; D008627:Mercuric Chloride; D008359:Mannose-6-Phosphate Isomerase; D003545:Cysteine