title: [Application of a model for gene local shuffling in ubiA mutation].
abstract: As a transhydrogen in the mitochondrial respiratory chain, Coenzyme Q (CoQ) has a critical role in the metabolism. 4-hydroxybenzoate polyprenyltransferase (UbiA), which coded by ubiA gene, is the rate-limiting enzyme in the CoQ biosynthesis progress. However, the relationship between structure and function remains unclear.
  
  
  OBJECTIVE
  To set up a synthetic oligonucleotide-based shuffling model and apply in the random mutation of DNA sequence of ubiA gene which codes putative active site.
  
  
  METHODS
  By using ubiA knockout mutant of E. coli. MC4100 as receptor, we set up a local Shuffling model by replacing target sequence with a random sequence fragment. Sequences of mutants and their function were analyzed.
  
  
  RESULTS
  After local Shuffling and two cycle of screening, we gained seven mutants. Compared to the wild type, most of the mutant amino sequences showed obvious change, and had different change trend of CoQ production. Sequencing result showed that three aspartic acid sites may be highly related to UbiA catalyze activity.
  
  
  CONCLUSIONS
  The local Shuffling model we set up is feasible. By applying this model, we preliminary verified several location of key amino acid sites of UbiA.
authors.affiliation: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, School of Biotechnology, Shanghai 200237, China. guchao_sh@yahoo.com.cn
authors.forename: Chao
authors.initials: C
authors.lastname: Gu
authors.forename: Nan
authors.initials: N
authors.lastname: Fu
authors.forename: Jiang
authors.initials: J
authors.lastname: Ye
authors.forename: Huizhan
authors.initials: H
authors.lastname: Zhang
mesh_terms: D000595:Amino Acid Sequence; D032621:DNA Shuffling / Q000379:methods*; D004122:Dimethylallyltranstransferase / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation
chemical_list: C509554:4-hydroxybenzoic acid oligoprenyltransferase, E coli; D004122:Dimethylallyltranstransferase