Purification and some properties of uridine diphosphate N-acetylglucosamine pyrophosphorylase from Neurospora crassa

Canadian Journal of Microbiology
K YamamotoT Tochikura

Abstract

Uridine diphosphate N-acetylglucosamine pyrophosphorylase (EC. 2.7.7.23) of Neurospora crassa has been purified approximately 210-fold with dithiothreitol as the stabilizing agent by use of chromatographic techniques. The enzyme preparation appeared to be homogeneous when subjected to electrophoresis. The molecular weight was estimated as approximately 37 000 by gel filtration. The enzyme had an isoelectric point around pH 4.4. Maximum activity of the enzyme was observed at pH 7.5. The enzyme required Mg2+, which may be replaced by other divalent cations such as Mn2+ and Co2+ for lesser degrees of effectiveness. The enzyme was strictly specific for UDP-N-acetylglucosamine as the substrate. The estimated values of Km were 2.2 mM for UDP-N-acetylglucosamine and 5.4 mM for inorganic pyrophosphate. The enzyme activity was highly stimulated by the addition of dithiothreitol or dithioerythritol but was lost by sulfhydryl inhibitory reagents.

Related Concepts

Neurospora crassa
Cations, Divalent
Neurospora
Dithiothreitol
Sulfhydryl Compounds
Sulfhydryl Reagents
Enzyme agent
Gel Chromatography
Enzyme Activity
Substrate Specificity

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