Studies of Streptomyces reticuli cel-1 (cellulase) gene expression in Streptomyces strains, Escherichia coli, and Bacillus subtilis

Abstract

Various Streptomyces strains [Streptomyces lividans 66, Streptomyces vinaceus, and Streptomyces coelicolor A3 (2)] acquired the ability to utilize crystalline cellulose (Avicel) after transformation with a multicopy vector containing the cel-1 gene from Streptomyces reticuli. The expression level in these hosts was two to three times lower than in S. reticuli, indicating the absence of positive regulatory elements. Like S. reticuli, they processed the Avicelase to its catalytic domain and to an enzymatically inactive part. The cel-1 gene with its original upstream region was not expressed within Escherichia coli. When cel-1 had been fused in phase with the lacZ gene, large quantities of the fusion protein were produced in E. coli. However, this protein was enzymatically inactive and proteolytically degraded to a series of truncated forms. As the cellulase (Avicelase) synthesized by S. reticuli is not cleaved by the E. coli proteases, its posttranslational modification is proposed. With Bacillus subtilis as host, the cel-1 gene was expressed neither under its own promoter nor under the control of a strong Bacillus promoter.

Binding modules alter the activity of chimeric cellulases: Effects of biomass pretreatment and enzyme source

“…Efforts to improve the catalytic activity of cellulases for depolymerization of biomass saccharides rely on production of cellulase variants for activity screening against solid substrates. Expressing cellulase variants in bacterial or yeast hosts is time‐consuming, can be cellulase specific (Walter and Schrempf, 1995), and often leads to inactive forms or low yields (Okada et al, 2000; Wang et al, 1994). These limitations have been obstacles for improving cellulases in a high‐throughput manner.…”

The Cell Wall-Anchored Streptomyces reticuli Avicel-Binding Protein (AbpS) and Its Gene

“…Physiological studies showed that Avicel, to which S. reticuli strongly adheres during cultivation (26), is the only known inducing carbon source (40). A low-molecular-weight inducer, such as the breakdown products of cellulose (glucose and cellobiose, -triose, -tetraose, and -pentaose), able to enter mycelia could be excluded (39). On the molecular level, the regulation of the S. reticuli cel1 gene is achieved by both transcriptional activation and repression.…”

Physiological Studies of Cellulase (Avicelase) Synthesis in Streptomyces reticuli

“…Several Streptomyces strains acquire the ability to degrade crystalline cellulose if they are transformed with the plasmid pZV1. This plasmid consists of a bifunctional vector (pZG6) into which the cel1 gene and its upstream region were inserted (13).…”

“…Expression of the cel1 gene in Streptomyces coelicolor and its glucose kinase-negative mutant M480. The plasmid pZV1 (13) was transformed in S. coelicolor M145 and S. coelicolor M480, a glucose kinase mutant of M145 (1). Unlike the progenitor strain (13), the transformants synthesized Avicelase activity in MM-3 medium containing yeast extract as the C source and Avicel as the inducer for cel1 expression.…”

“…The plasmid pZV1 (13) was transformed in S. coelicolor M145 and S. coelicolor M480, a glucose kinase mutant of M145 (1). Unlike the progenitor strain (13), the transformants synthesized Avicelase activity in MM-3 medium containing yeast extract as the C source and Avicel as the inducer for cel1 expression. The addition of glucose and glycerol led to a significant repression of Avicelase synthesis in M145(pZV1).…”