Crystallization and preliminary crystallographic analysis of poly(3-hydroxybutyrate) depolymerase fromBacillus thuringiensis

Abstract

Poly[(R)-3-hydroxybutyrate] (PHB) is a microbial biopolymer that has been commercialized as biodegradable plastics. The key enzyme for the degradation is PHB depolymerase (PhaZ). A new intracellular PhaZ from Bacillus thuringiensis (BtPhaZ) has been screened for potential applications in polymer biodegradation. Recombinant BtPhaZ was crystallized using 25% polyethylene glycol 3350, 0.2 M ammonium acetate, 0.1 M bis-tris pH 6.5 at 288 K. The crystals belonged to space group P1, with unit-cell parameters a = 42.97, b = 83.23, c = 85.50 Å , = 73.45, = 82.83, = 83.49 . An X-ray diffraction data set was collected to 1.42 Å resolution with an R merge of 6.4%. Unexpectedly, a molecular-replacement solution was obtained using the crystal structure of Streptomyces lividans chloroperoxidase as a template, which shares 24% sequence identity to BtPhaZ. This is the first crystal structure of an intracellular poly(3-hydroxybutyrate) depolymerase.

Poly-β-hydroxybutyrate Metabolism Is Unrelated to the Sporulation and Parasporal Crystal Protein Formation in Bacillus thuringiensis

“…Most methods to identify new nPHB depolymerase is based on sequence comparison with already known depolymerase, either by comparing their phaZ genes in the other species, or phaC genes ( Tseng et al, 2006 ; Chen et al, 2009 ; Sznajder and Jendrossek, 2014 ), or through the blast search of the consensus motif. To date, only one PHB degradation enzyme (PhaZ) was well characterized in B. thuringiensis ( Tseng et al, 2006 ; Wang et al, 2014 ). Obtained results indicated that these methods all suffer from some limitations.…”