The pla2 gene encoding a phospholipase A(2) (EC 3.1.1.4) of Lactobacillus casei DSM20011 was cloned and expressed in the yeast Kluyveromyces lactis GG799 successfully for the first time. The structural pla2 gene fused in frame with the K. lactis secretion signal alpha-mating factor was integrated into the LAC4 locus and expressed under the control of the LAC4 promoter. sPLA(2) activity was detected in the culture supernatant during shake flask culture of K. lactis/pKLAC1-pla2. In comparison with the control strain K. lactis/pKLAC1, SDS-PAGE analysis revealed a 17-kDa recombinant protein band in K. lactis/pKLAC1-pla2, which was consistent with the predicted molecular weight of the mature protein. Real-time quantitative PCR analysis indicated that the copy number of the integrated pla2 gene ranged from 2 to 6 and positively correlated with sPLA(2) activity. When the inducer galactose was used as the carbon source, the sPLA(2) activity in the culture supernatant of the recombinant that harbored six pla2 gene copies reached 1.96 +/- 0.15 U/mL. The influence of the culture composition and conditions on the recombinant sPLA(2) activity in shake flask culture were also studied. When the recombinant was cultured at 30 degrees C in a YPD medium culture volume of 70 mL in a 250-mL shake flask with an initial pH of 7.0, the sPLA(2) activity reached 2.16 +/- 0.18 U/mL. (c) 2015, The Society for Biotechnology, Japan. All rights reserved.