BACKGROUND & AIMS:
:The thermophilic actinomycete Thermomonospora fusca produced endoxylanase, alpha-arabinofuranosidase, beta-xylosidase, and acetyl esterase activities maximally during growth on xylan. Growth yields on glucose, xylose, or arabinose were comparable, but production of endoxylanase and beta-xylosidase was not induced on these substrates. The crude xylanase activity was thermostable and relatively resistant to end product inhibition by xylobiose and xylan hydrolysis products. Six proteins with xylanase activity were identified by zymogram analysis of isoelectric focusing gels, but only a 32-kDa protein exhibiting three isomeric forms could be purified by fast protein liquid chromatography. Endoglucanases were also identified in carboxymethylcellulose-grown cultures, and their distinction from endoxylanases was confirmed. alpha-Arabinofuranosidase activity was due to a single dimeric protein of 92 kDa, which was particularly resistant to end product inhibition by arabinose. Three bands of acetyl esterase activity were detected by zymogram analysis, and there was evidence that these mainly consisted of an intracellular 80-kDa protein secreted to yield active 40-kDa subunits in the culture supernatant. The acetyl esterases were found to be responsible for acetyl xylan esterase activity in T. fusca, in contrast to the distinction proposed in some other systems. The addition of purified betaxylosidase to endoxylanase increased the hydrolysis of xylan, probably by relieving end product inhibition. The enhanced saccharification of wheat straw caused by the addition of purified alpha-arabinofuranosidase to T. fusca endoxylanase suggested a truly synergistic relationship, in agreement with proposals that arabinose side groups on the xylan chain participate in cross-linking within the plant cell wall structure.
背景与目标:
: 嗜热放线菌Thermomonospora fusca在木聚糖上生长过程中最大程度地产生木聚糖酶,α-阿拉伯呋喃糖苷酶,β-木糖苷酶和乙酰酯酶活性。葡萄糖,木糖或阿拉伯糖的生长产量相当,但在这些底物上未诱导内切木聚糖酶和 β-木糖苷酶的产生。粗木聚糖酶活性是热稳定性的,并且相对抵抗木糖和木聚糖水解产物对终产物的抑制。通过等电聚焦凝胶的酶谱分析鉴定了六种具有木聚糖酶活性的蛋白质,但是通过快速蛋白质液相色谱法只能纯化表现出三种异构体形式的32 kDa蛋白质。在羧甲基纤维素生长的培养物中也鉴定了内切葡聚糖酶,并确认了它们与内切木聚糖酶的区别。Α-阿拉伯呋喃糖苷酶的活性归因于92 kDa的单个二聚体蛋白,该蛋白特别能抵抗阿拉伯糖的终产物抑制。通过酶谱分析检测到三个乙酰酯酶活性带,有证据表明,这些带主要由分泌的细胞内80 kDa蛋白组成,在培养上清液中产生活性40 kDa亚基。与其他一些系统中提出的区别相反,发现乙酰酯酶负责T. fusca中的乙酰木聚糖酯酶活性。在木聚糖内切糖苷酶中添加纯化的甜蜜糖糖苷酶可能通过减轻终产物的抑制作用而增加了木聚糖的水解。通过向T. fusca内切木聚糖酶中添加纯化的 α-阿拉伯呋喃糖苷酶引起的小麦秸秆糖化作用增强,这与木聚糖链上的阿拉伯糖侧基参与植物细胞壁内交联的建议一致结构。