title: Novel and efficient screening of PQQ high-yielding strains and subsequent cultivation optimization.
abstract: A novel and efficient screening method for pyrroloquinoline quinone (PQQ) high-yielding methylotrophic strains was developed by using glucose dehydrogenase apoenzyme (GDHA) which depended on PQQ as the cofactor. Using this high-throughput method, PQQ high-yielding strains were rapidly screened out from thousands of methylotrophic colonies at a time. The comprehensive phylogenetic analysis revealed that the highest PQQ-producing strain zju323 (CCTCC M 2016079) could be assigned to a novel species in the genus Methylobacillus of the Betaproteobacteria. After systematic optimization of different medium components and cultivation conditions, about 33.4 mg/L of PQQ was obtained after 48 h of cultivation with Methylobacillus sp. zju323 at the shake flask scale. Further cultivations of Methylobacillus sp. zju323 were carried out to investigate the biosynthesis of PQQ in 10-L bench-top fermenters. In the batch operation, the PQQ accumulation reached 78 mg/L in the broth after 53 h of cultivation. By adopting methanol feeding strategy, the highest PQQ concentration was improved up to 162.2 mg/L after 75 h of cultivation. This work developed a high-throughput strategy of screening PQQ-producing strains from soil samples and also demonstrated one potential bioprocess for large-scale PQQ production with the isolated PQQ strain.
authors.affiliation: Key Laboratory of Biomass Chemical Engineering (Ministry of Education), College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, People's Republic of China.|Institute of Biological Engineering, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, People's Republic of China.
authors.forename: Zhenjun
authors.initials: Z
authors.lastname: Si
authors.affiliation: Institute of Biological Engineering, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, People's Republic of China.
authors.forename: Jianzhong
authors.initials: J
authors.lastname: Zhu
authors.affiliation: Institute of Biological Engineering, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, People's Republic of China.
authors.forename: Wenguang
authors.initials: W
authors.lastname: Wang
authors.affiliation: Key Laboratory of Biomass Chemical Engineering (Ministry of Education), College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, People's Republic of China.|Institute of Biological Engineering, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, People's Republic of China.
authors.forename: Lei
authors.initials: L
authors.lastname: Huang
authors.affiliation: School of Biological and Chemical Engineering, Zhejiang University of Science and Technology, Hangzhou, 310023, People's Republic of China. weipeilian@126.com.
authors.forename: Peilian
authors.initials: P
authors.lastname: Wei
authors.affiliation: Key Laboratory of Biomass Chemical Engineering (Ministry of Education), College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, People's Republic of China.
authors.forename: Jin
authors.initials: J
authors.lastname: Cai
authors.affiliation: Key Laboratory of Biomass Chemical Engineering (Ministry of Education), College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, People's Republic of China. znxu@zju.edu.cn.|Institute of Biological Engineering, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou, 310027, People's Republic of China. znxu@zju.edu.cn.
authors.forename: Zhinan
authors.initials: Z
authors.lastname: Xu
mesh_terms: D003470:Culture Media / Q000737:chemistry; D005285:Fermentation; D005948:Glucose Dehydrogenases / Q000378:metabolism; D008403:Mass Screening / Q000379:methods*; D020631:Methylobacillus / Q000145:classification / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism*; D008828:Microbiological Techniques / Q000379:methods; D045542:PQQ Cofactor / Q000378:metabolism*; D010802:Phylogeny
chemical_list: D003470:Culture Media; D045542:PQQ Cofactor; D005948:Glucose Dehydrogenases; C033955:glucose dehydrogenase (pyrroloquinoline-quinone)
keywords: Fed-batch fermentation; High-throughput screening; Methylobacillus; PQQ