Molecular identification of the industrially important strain Ogataea parapolymorpha

Наумова, Е. С.; Dmitruk, K V; Kshanovskaya, B. V.; Sibirny, Andriy А.; Наумов, Г. И.

doi:10.1134/s0026261713030090

Cited by 3 publications

(1 citation statement)

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“…The strain is isogenic to CBS4732 (Lahtchev et al , ), which was earlier allocated to the species Hansenula polymorpha . Several sibling species in the H. polymorpha complex were described earlier (Naumov et al ., ) and currently (Suh and Zhou, ; Naumova et al ., ) the CBS4732 strain belongs to Ogataea polymorpha . Op wild‐type strain and respective deletion mutants of maltase and α ‐glucoside permease genes (Alamäe et al , ; Viigand and Alamäe, ) were grown in 0.67% yeast nitrogen base (YNB) medium (Difco, USA) without amino acids, with required auxotrophic supplements added; 2% agar was used for solid media.…”

Section: Methodsmentioning

confidence: 99%

Maltase protein of Ogataea (Hansenula) polymorpha is a counterpart to the resurrected ancestor protein ancMALS of yeast maltases and isomaltases

Viigand

Visnapuu

Mardo

et al. 2016

Yeast

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Saccharomyces cerevisiae maltases use maltose, maltulose, turanose and maltotriose as substrates, isomaltases use isomaltose, α‐methylglucoside and palatinose and both use sucrose. These enzymes are hypothesized to have evolved from a promiscuous α‐glucosidase ancMALS through duplication and mutation of the genes. We studied substrate specificity of the maltase protein MAL1 from an earlier diverged yeast, Ogataea polymorpha (Op), in the light of this hypothesis. MAL1 has extended substrate specificity and its properties are strikingly similar to those of resurrected ancMALS. Moreover, amino acids considered to determine selective substrate binding are highly conserved between Op MAL1 and ancMALS. Op MAL1 represents an α‐glucosidase in which both maltase and isomaltase activities are well optimized in a single enzyme. Substitution of Thr200 (corresponds to Val216 in S. cerevisiae isomaltase IMA1) with Val in MAL1 drastically reduced the hydrolysis of maltose‐like substrates (α‐1,4‐glucosides), confirming the requirement of Thr at the respective position for this function. Differential scanning fluorimetry (DSF) of the catalytically inactive mutant Asp199Ala of MAL1 in the presence of its substrates and selected monosaccharides suggested that the substrate‐binding pocket of MAL1 has three subsites (–1, +1 and +2) and that binding is strongest at the –1 subsite. The DSF assay results were in good accordance with affinity (K m) and inhibition (K i) data of the enzyme for tested substrates, indicating the power of the method to predict substrate binding. Deletion of either the maltase (MAL1) or α‐glucoside permease (MAL2) gene in Op abolished the growth of yeast on MAL1 substrates, confirming the requirement of both proteins for usage of these sugars. © 2016 The Authors. Yeast published by John Wiley & Sons, Ltd.

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