Institute of Microbial Technology, Council of Scientific and Industrial Research, Sector 39A, Chandigarh 160 036, India.
Appl Microbiol Biotechnol. 2010 Dec;88(6):1311-20. doi: 10.1007/s00253-010-2818-6. Epub 2010 Aug 18.
Pichia angusta (syn. Hansenula polymorpha) represents one of the rare yeast that can grow and ferment both xylose and glucose at higher temperature (50°C). However, little is known about the enzymes involved in xylose utilization from this species. Previous studies indicated the presence of one xylose reductase and two xylitol dehydrogenase genes in P. angusta. In this study, we have expressed both xylitol dehydrogenases (PaXdh1p and PaXdh2p) in Escherichia coli and purified them as 6X-Histidine-tagged proteins. Biochemical characterization of the recombinant proteins reveals that both PaXdh1p and PaXdh2p are thermotolerant enzymes. PaXdh2p contains a catalytic and a structural Zn atom. However, the structural Zn atom is not present in PaXdh1p. Both enzymes also differ in their affinity for the substrate as well as in the catalytic efficiency. Through mutagenesis and modeling approaches, we have also identified residues important for catalysis and substrate binding.
威克汉姆酵母（syn. Hansenula polymorpha）是少数能够在较高温度（50°C）下同时利用木糖和葡萄糖进行生长和发酵的酵母之一。然而，对于该物种中参与木糖利用的酶知之甚少。先前的研究表明，威克汉姆酵母中存在一个木糖还原酶和两个木酮糖脱氢酶基因。在本研究中，我们在大肠杆菌中表达了两种木酮糖脱氢酶（PaXdh1p 和 PaXdh2p），并将其纯化为 6X-His 标签蛋白。对重组蛋白的生化特性分析表明，PaXdh1p 和 PaXdh2p 都是耐热酶。PaXdh2p 含有一个催化和一个结构 Zn 原子。然而，结构 Zn 原子不存在于 PaXdh1p 中。两种酶在底物亲和力和催化效率上也存在差异。通过突变和建模方法，我们还确定了催化和底物结合的重要残基。
