MOLECULAR ADAPTATION OF DROSOPHILA-MELANOGASTER LYSOZYMES TO A DIGESTIVE FUNCTION

A lysozyme (pI 5.5) was purified to homogeneity from heated acid extra cts of Drosophila melanogaster larvae, using gel filtration in a Super ose column and ion-exchange chromatography in a Mono Q column. The fin al yield was 67%. The purified lysozyme with Mr 13 700 (determined by SDS-polyacrylamide gel electrophoresis) decreases in activity and has its pH optimum displaced towards acidic values and Kin increases as th e ionic strength of the medium becomes higher. The lysozyme is resista nt to a cathepsin D-like proteinase present in cyclorrhaphous Diptera and displays a chitinase activity which is 11-fold higher than that of chicken lysozyme. Microsequencing of an internal peptide of the purif ied lysozyme showed that this enzyme is the product of the previously sequenced Lys D gene. The results suggest that the product of the Lys P gene has pI 7.2, a pH optimum around 5 and is not a true digestive e nzyme. The most remarkable sequence convergence of D, melanogaster lys ozyme D and lysozymes from vertebrate foregut fermenters are serine 10 4 and a decrease in the number of basic amino acids, suggesting that t hese features are necessary for digestive function in an acid environm ent. Adaptive residues putatively conferring stability in an acid prot eolytic environment differ between insects and vertebrates, probably b ecause they depend on the overall three-dimensional structure of the l ysozymes. A maximum likelihood phylogeny and inferences from insect ly sozyme sequences showed that the recruitment of lysozymes as digestive enzymes is an ancestral condition of the flies (Diptera: Cyclorrhapha ). (C) 1998 Elsevier Science Ltd. All rights reserved.