Part:BBa_K3075001

DBAT^G38R/F301V-SnoopT-His

Introduction

DBAT-G38R/F301V-SnoopT consists of the enzyme 10-deacetylbaccatin III 10-O-acetyltransferase (DBAT) fused at the C-terminus to a short polypeptide tag (Snooptag) and a Hexahistidine Tag (6xHis-tag), separated by interconnecting GSG linkage sequences. The sequence of DBAT which was used, originated from Taxus cuspidata (Japanese yew), but contained a double mutation of G38R/F301V. (2)

Usage and Biology

This protein naturally participates in the synthesis of baccatin III from 10-deacetyl-2-debenzoylbaccatin III, where it catalyses the final acetylation of. Baccatin III synthesis is a subpathway of paclitaxel biosynthesis, which is itself part of Alkaloid biosynthesis. The mutant however, has been designed to catalyse the acetylation of 10-deacetyltaxol (DT) with a catalytic efficiency approximately six times higher than that of the wild-type. (2) The recombinant mutant enzyme has a length of 440 amino acid residues, a molecular weight of 49,052 Da and an optimum pH of 7.5. (3)

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal PstI site found at 771
  Illegal PstI site found at 826
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal PstI site found at 771
  Illegal PstI site found at 826
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 28
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal PstI site found at 771
  Illegal PstI site found at 826
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal PstI site found at 771
  Illegal PstI site found at 826
  
• 1000
  COMPATIBLE WITH RFC[1000]