title: Acthi, a thiazole biosynthesis enzyme, is essential for thiamine biosynthesis and CPC production in Acremonium chrysogenum.
abstract: BACKGROUND
  The filamentous fungus Acremonium chrysogenum is an important industrial fungus and is used in the production of the β-lactam antibiotic cephalosporin C. Little is known regarding the molecular and biological mechanisms of how this industrial strain was improved by mutagenesis and molecular breeding. Comparative proteomics is one of the most powerful methods to evaluate the influence of gene expression on metabolite production.
  
  
  RESULTS
  In this study, we used comparative proteomics to investigate the molecular mechanisms involved in the biosynthesis of cephalosporin C between a high-producer (HY) strain and a wide-type (WT) strain. We found that the expression levels of thiamine biosynthesis-related enzymes, including the thiazole biosynthesis enzyme (Acthi), pyruvate oxidase, flavin adenine dinucleotide (FAD)-dependent oxidoreductase and sulfur carrier protein-thiS, were up-regulated in the HY strain. An Acthi-silencing mutant of the WT strain grew poorly on chemically defined medium (MMC) in the absence of thiamine, and its growth was recovered on MMC medium supplemented with thiamine. The intracellular thiamine content was changed in the Acthi silencing or over-expression mutants. In addition, we demonstrated that the manipulation of the Acthi gene can affect the hyphal growth of Acremonium chrysogenum, the transcription levels of cephalosporin C biosynthetic genes, the quantification levels of precursor amino acids for cephalosporin C synthesis and the expression levels of thiamine diphosphate-dependent enzymes. Over-expression of Acthi can significantly increase the cephalosporin C yield in both the WT strain and the HY mutant strain.
  
  
  CONCLUSIONS
  Using comparative proteomics, four differently expressed proteins were exploited, whose functions may be involved in thiamine diphosphate metabolism. Among these proteins, the thiazole biosynthesis enzyme (ActhiS) may play an important role in cephalosporin C biosynthesis. Our studies suggested that Acthi might be involved in the transcriptional regulation of cephalosporin C biosynthesis. Therefore, the thiamine metabolic pathway could be a potential target for the molecular breeding of this cephalosporin C producer for industrial applications.
authors.affiliation: China State Institute of Pharmaceutical Industry, Zhangjiang Institute, 1599 Zhangheng Road, Shanghai, 201203, China. yanzi19831221@126.com.|Shanghai Institute of Pharmaceutical Industry, 1320 Beijing Road (W), Shanghai, 200040, China. yanzi19831221@126.com.
authors.forename: Yan
authors.initials: Y
authors.lastname: Liu
authors.affiliation: China State Institute of Pharmaceutical Industry, Zhangjiang Institute, 1599 Zhangheng Road, Shanghai, 201203, China. zhangwei@sinopharm.com.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: China State Institute of Pharmaceutical Industry, Zhangjiang Institute, 1599 Zhangheng Road, Shanghai, 201203, China. xieliping@sinopharm.com.
authors.forename: Liping
authors.initials: L
authors.lastname: Xie
authors.affiliation: Shanghai Institute of Pharmaceutical Industry, 1320 Beijing Road (W), Shanghai, 200040, China. liuhongrd@126.com.|Present address: Luye Pharma Group Ltd., Yantai, Shandong, 264003, China. liuhongrd@126.com.
authors.forename: Hong
authors.initials: H
authors.lastname: Liu
authors.affiliation: China State Institute of Pharmaceutical Industry, Zhangjiang Institute, 1599 Zhangheng Road, Shanghai, 201203, China. gongguihua@sinopharm.com.
authors.forename: Guihua
authors.initials: G
authors.lastname: Gong
authors.affiliation: Shanghai Institute of Pharmaceutical Industry, 1320 Beijing Road (W), Shanghai, 200040, China. baoquanzhu@sipi.com.cn.
authors.forename: Baoquan
authors.initials: B
authors.lastname: Zhu
authors.affiliation: China State Institute of Pharmaceutical Industry, Zhangjiang Institute, 1599 Zhangheng Road, Shanghai, 201203, China. bebydou@hotmail.com.
authors.forename: Youjia
authors.initials: Y
authors.lastname: Hu
mesh_terms: D000164:Acremonium / Q000201:enzymology*; D002511:Cephalosporins / Q000378:metabolism*; D014176:Protein Biosynthesis; D013831:Thiamine / Q000096:biosynthesis / Q000378:metabolism*; D013844:Thiazoles / Q000378:metabolism*
chemical_list: D002511:Cephalosporins; D013844:Thiazoles; C025163:cephalosporin C; D013831:Thiamine