Human non-pancreatic secretory phospholipase A(2) (hnpsPLA(2)) catalyzes the sn-2 acyl hydrolysis of phospholipids. It was reported that hnpsPLA(2) is involved in various diseases like inflammation, cancer, and so on. This enzyme also exhibits anti-bacterial and anti-virus activities. It is active over a broad pH range, with higher activity at alkaline conditions. In order to make it suitable as a potential bactericide, high activity at neutral pH is preferable. We have tried to tailor the pH dependence of hnpsPLA(2) activity by replacing its surface charged residues. Three surface charge replacements, Arg42Glu, Arg100Glu, and Glu89Lys, showed increased activities at neutral pH, which are 2.3, 2.8, and 2.3 times that of the wild-type enzyme at pH 7. Both the positive-to-negative and negative-to-positive mutations lowered the optimum enzymatic reaction pH of hnpsPLA(2), indicating that the enzyme pH profile depends on a delicate balance of charged residues. The activity changes are in good agreement with the recently proposed calcium-coordinated catalytic triad mechanism. This study also provides a general means of enhancing hnpsPLA(2) activity at low pH.