Alcocer M J C, Furniss C S M, Kroon P A, Campbell M, Archer D B
School of Life and Environmental Sciences, University of Nottingham, University Park, NG7 2RD, Nottingham,
Appl Microbiol Biotechnol. 2003 Feb;60(6):726-32. doi: 10.1007/s00253-002-1184-4. Epub 2002 Dec 21.
Genes encoding three enzymes with xylanase activity from the filamentous fungus Penicillium funiculosum are described. Two of the encoded xylanases are predicted to be modular in structure with catalytic and substrate-binding domains separated by a serine and threonine-rich linker region; the other had none of these properties and was non-modular. In order to develop P. funiculosum as a host for the secreted production of heterologous proteins, each of the xylanases was assessed for use as a carrier protein in a fusion strategy. We show that one of the modular xylanases (encoded by xynA) was an effective carrier protein but the other (encoded by xynB) and the non-modular xylanase (encoded by xynC) were not effective as secretion carriers. We show that the beta-glucuronidase (GUS) protein from Escherichia coli is secreted by P. funiculosum when expressed as an XYNA fusion but that the secreted GUS protein, cleaved in vivo from XYNA, is glycosylated and enzymatically inactive.
本文描述了来自丝状真菌绳状青霉的三种具有木聚糖酶活性的酶的编码基因。其中两种编码的木聚糖酶预计结构为模块化，催化结构域和底物结合结构域由富含丝氨酸和苏氨酸的连接区隔开；另一种则没有这些特性，是非模块化的。为了将绳状青霉开发成用于分泌生产异源蛋白的宿主，对每种木聚糖酶进行了评估，以确定其在融合策略中作为载体蛋白的用途。我们发现，其中一种模块化木聚糖酶（由xynA编码）是一种有效的载体蛋白，但另一种（由xynB编码）和非模块化木聚糖酶（由xynC编码）作为分泌载体无效。我们还发现，当大肠杆菌的β-葡萄糖醛酸酶（GUS）蛋白作为XYNA融合蛋白表达时，可由绳状青霉分泌，但从XYNA体内切割下来的分泌型GUS蛋白会发生糖基化且无酶活性。
