Secretory overexpression of heterologous fungal cellulase genes in Lipomyces starkeyi

Lipomyces starkeyi is one of the highest lipid-producing microorganisms characterized to date. To investigate the possibility of producing lipid or fatty acid-related biofuels directly from abundant and low-cost lignocellulosic substrates using L. starkeyi, we investigated its ability for overexpressing and secreting key fungal cellulase genes.  A total of four customized signal peptides, including three from native L. starkeyi and one from Yarrowia lipolytica, were selected and investigated as guides for the secretion of each of two recombinant cellulase proteins, which are a native Trichoderma reesei endoglucanase-II (EGII) and a chimeric cellobiohydrolase (CBH) formed by fusion of the catalytic domain from Talaromyces emersonii Cel7A with the linker and cellulose-binding domain of T. reesei Cel7A. We found that all four signal peptides worked for secretion of the EGII protein, whereas only three of them have as yet been shown to work for the chimeric CBH. The recombinant CBH proteins were almost completely secreted into medium, indicating high efficiency of protein secretion guided by the signal peptide. Yield of the recombinant EGII protein was much higher than that of the recombinant CBH when the same signal peptides were employed. Surprisingly, one Y. lipolytica signal peptide was able to guide the secretion of recombinant CBH protein from L. starkeyi as efficiently as did the native L. starkeyi signal peptides. The purified recombinant CBHI showed high activity against pretreated corn stover, and purified recombinant T. reesei EGII showed high endocellulase activity measured by the CELLG3 method.