Mutational analysis of human lipoprotein lipase by carboxy-terminal truncation

Abstract

We have previously reported a Trp382 (TGG)–>stop (TGA) mutation that causes familial lipoprotein lipase (LPL) deficiency. Expression study of the Trp382–>stop mutant showed that the truncated LPL was catalytically inactive with a marked reduction in the expressed mass. To investigate the minimal amino-terminal sequence of human LPL required for an active enzyme, a series of carboxy-terminal (C-terminal) truncated LPLs were expressed in vitro, and the enzyme activity, mass, and LPL mRNA levels were analyzed. The lipolytic activity showed a stepwise reduction between LPL-437 (Cys438–>stop; 68% of normal LPL activity in medium) and LPL-434 (Phe435–>stop; 3%). Without affecting LPL mRNA levels, LPL mass was reduced with the mutants not larger than LPL-437. In terms of specific activity, a significant difference was observed between LPL-436 (Lys437–>stop; 88% of that of normal LPL in medium) and LPL-435 (Val436–>stop; 22%), implying the importance of the role of Val436 in LPL action. Furthermore, our results unexpectedly showed that LPL-446 (Ser447–>stop), which is considered to be a common polymorphic form of LPL, exhibited higher activity than normal LPL (185% in medium). These results demonstrate that the C-terminal region of human LPL is closely associated with the expression of enzyme mass and catalytic activity.