title: Secretory pathway engineering enhances secretion of cellobiohydrolase I from Trichoderma reesei in Saccharomyces cerevisiae.
abstract: Improving the cellulase secretion is beneficial for Saccharomyces cerevisiae used in consolidated bioprocessing (CBP) of cellulosic ethanol. In this study, protein secretory pathway, including protein folding, disulfide bond formation, and protein trafficking and sorting, was modified in S. cerevisiae. The effects of these modifications on the secretion of cellobiohydrolase I (Tr-Cel7A) with its native signal peptide from Trichoderma reesei were investigated. The results showed that overexpression of the protein disulfide isomerase Sc-PDI1 and the plasma membrane targeting soluble N-ethylmaleimide-sensitive factor attachment protein receptor Sc-SSO1, and disruption of the sorting receptor Sc-VPS10 and a Ca(2+)/Mn(2+) ATPase Sc-PMR1, improved respectively the extracellular Tr-Cel7A activities. Among them, disruption of Sc-PMR1 showed better improvement of 162% in the extracellular activity and decreased the glycosylation of Tr-Cel7A. Multiple modifications generally resulted in higher activities. The extracellular activities of the quadruple-modified strain (vps10Δ/pmr1Δ/SSO1/PDI1/cel7AF) using p-nitrophenyl-β-d-cellobioside (pNPC) and phosphoric acid swollen cellulose (PASC) as the substrates, respectively, were 3.9-fold and 1.3-fold higher than that of the reference strain cel7AF. The results indicated that engineering of the protein secretory pathway is an effective approach to improve the Tr-Cel7A secretion in S. cerevisiae.
authors.affiliation: The State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China. Electronic address: lili_xu@126.com.
authors.forename: Lili
authors.initials: L
authors.lastname: Xu
authors.affiliation: The State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China. Electronic address: shenyu@sdu.edu.cn.
authors.forename: Yu
authors.initials: Y
authors.lastname: Shen
authors.affiliation: The State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China. Electronic address: houjin@sdu.edu.cn.
authors.forename: Jin
authors.initials: J
authors.lastname: Hou
authors.affiliation: The State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China. Electronic address: pengbingyin@yahoo.com.cn.
authors.forename: Bingyin
authors.initials: B
authors.lastname: Peng
authors.affiliation: The State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China. Electronic address: hongtingtang@126.com.
authors.forename: Hongting
authors.initials: H
authors.lastname: Tang
authors.affiliation: The State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China. Electronic address: bxm@sdu.edu.cn.
authors.forename: Xiaoming
authors.initials: X
authors.lastname: Bao
mesh_terms: D015153:Blotting, Western; D000252:Calcium-Transporting ATPases / Q000235:genetics / Q000378:metabolism; D002482:Cellulose / Q000378:metabolism*; D043366:Cellulose 1,4-beta-Cellobiosidase / Q000378:metabolism*; D004220:Disulfides / Q000378:metabolism; D000431:Ethanol / Q000378:metabolism; D006031:Glycosylation; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism; D019704:Protein Disulfide-Isomerases / Q000235:genetics / Q000378:metabolism; D017510:Protein Folding; D011499:Protein Processing, Post-Translational; D021381:Protein Transport; D050765:Qa-SNARE Proteins / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D055571:Secretory Pathway / Q000502:physiology*; D013379:Substrate Specificity; D014242:Trichoderma / Q000201:enzymology*; D033921:Vesicular Transport Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D004220:Disulfides; D018832:Molecular Chaperones; C080576:PDI1 protein, S cerevisiae; C087335:PEP1 protein, S cerevisiae; D050765:Qa-SNARE Proteins; D012333:RNA, Messenger; C101767:SSC1 protein, S cerevisiae; C084006:SSO1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D033921:Vesicular Transport Proteins; D000431:Ethanol; D002482:Cellulose; D043366:Cellulose 1,4-beta-Cellobiosidase; D019704:Protein Disulfide-Isomerases; D000252:Calcium-Transporting ATPases
keywords: Budding yeast; Cel7A; Cellulosic ethanol; Consolidated bioprocessing; PMR1; Protein secretion