Characterization of an arylsulfatase from a mutant library of Pseudoalteromonas carrageenovora arylsulfatase

- A library of Pseudoalteromonas carrageenovora arylsulfatase mutants was constructed based on error-prone PCR method. We obtained a mutant arylsulfatase H260L with improved thermal stability. This mutant arylsulfatase exhibited stability across a pH range of 6.0-9.0. It had a relatively good resistance against some detected detergents. We identified the desulfation activity of H260L for the sulfate groups in Gracilaria lemaneiformis agar.

A library of Pseudoalteromonas carrageenovora arylsulfatase mutants was constructed by introducing random mutagenesis using error-prone PCR. After screening, one mutant strain was obtained whose arylsulfatase had improved thermal stability. Protein sequence analysis revealed one amino acid substitution of H260L. The mutant arylsulfatase (named H260L) retained higher residual activity than wild-type enzyme (named WT) after incubation at 45, 50, 55 and 60 °C for 60 min. Thermal inactivation analysis showed that the half-life (t1/2) value at 55 °C for H260L was 40.6 min, while that of WT was 9.1 min. When p-nitrophenyl sulfate was used as a substrate, the optimal reaction temperature and pH for the mutant enzyme were 55 °C and pH 8.0, respectively. H260L was stable over the pH range of 6.0-9.0. Inhibition assay with EDTA indicated that metal ions play an important role during the catalytic process of the mutant enzyme. The desulfation ratio against agar of Gracilaria lemaneiformis was 82%.