Gene expression and characterization of a serine proteinase inhibitor PmSERPIN8 from the black tiger shrimp Penaeus monodon

The ubiquitous SERPINs or serine proteinase inhibitors are essential for controlling proteinases in several biological processes in various organisms. A PmSERPIN8, one of eight SERPINs identified from the Penaeus monodon database, is studied and reported herein. The open reading frame of PmSERPIN8 gene derived from a genomic gene contains 5 exons of 320, 139, 244, 239 and 312 bp separated by 4 introns of 447, 657, 326 and 479 bp. The PmSERPIN8 gene is highly expressed at nauplius stage and gradually subsided as the shrimp grow through zoea, mysis and postlarva stages. At sub-adult stage, the PmSERPIN8 gene is expressed mainly in the hemocyte and epipodite. The expression in response to Vibrio harveyi and YHV injection is up-regulated, respectively, at 24 and 48 h post-injection. The number of PmSERPIN8-producing hemocytes, however, is observed highest at 48 h post V. harveyi injection. All three hemocyte cell types: hyaline, semigranular and granular hemocytes are able to produce PmSERPIN8. The recombinant mature PmSERPIN8 (rPmSERPIN8) with a predicted size of 45.5 kDa was over-produced in an Escherichia coli system, solubilized from the inclusion bodies, purified and tested for its activity. We have found that the rPmSERPIN8 is able to inhibit the growth of Gram-positive bacterium, Bacillus subtilis, but not Gram-negative bacterium, V. harveyi 639, and inhibit the shrimp prophenoloxidase system. The PmSERPIN8 is, thus, involved in the shrimp innate immunity.