Characteristics of Thermo Tolerant Serine Peptidase Vpr from Endophytic
Bacillus cereus strain InaCC-B1657
Abstract
Bacterial endophytes are a promising source of novel fibrinolytic
enzymes with unique metabolic pathways and desirable characteristics
that may not be present from conventionally explored sources. This study
aimed to characterize fibrinolytic enzymes from selected endophytic
bacteria isolated from papaya (Carica papaya L.) leaves and the genes
encoding the enzymes. A strain BFP1 (InaCC-B1657) that showed the
highest fibrinolytic activity was identified as Bacillus cereus based on
the phylogenetic analysis of the 16S rRNA sequence. The enzyme exhibited
optimum activity at 50°C and pH 7.0, and remained stable until 80°C and
pH 6 - 10 for 24 h. The assay of metal ions and inhibitors on the
fibrinolytic enzyme activity found that adding Cu2+ stimulated, while
Fe2+ reduced the activity. PMSF and TPCK inhibited the enzyme activity,
while adding EDTA and EGTA increased the activity. These suggest that
the fibrinolytic enzymes belong to the serine protease group. Of the 21
proteases/peptidases determined from the 5,257,484 base pairs (bp)
genome, minor extracellular protease Vpr and S8 family peptidase genes
were found related to the fibrinolytic enzyme activity. The Vpr gene has
a molecular weight of 98.5 kDa. The subtilase domain (peptidase S8
family) and the catalytic triad subtilase active sites (Asp204, His237,
and Ser531) were detected. A prediction of physicochemical
characteristics of the Vpr gene showed that the enzyme is hydrophilic
and exhibited alkali-halo tolerant and thermo tolerant over a broad
range of temperatures.