MALTASE-GLUCOAMYLASE (MGA) MAY BE AN ALTERNATE PATHWAY FOR STARCH DIGESTION IN YOUNG INFANTS

Abstract

Mothers continue to feed cereals despite pediatric advice that, because of pancreatic and salivary amylase immaturity, starches not be fed to young infants. Because intestinal glucoamylase digests starch, we hypothesized that human MGA may serve as an important alternate starch digestion pathway. The report of a diet responsive form of chronic diarrhea in children with selective loss of mucosal starch hydrolyzing activity (Lebenthal, E. et al (1994) J. Pediatr.124, 541-546) supports this hypothesis. An overlap exists between in vitro activities of the mucosal glucohydrolase complexes which has prevented direct enzymatic testing of our hypothesis. Aims: The objective of our study was the cloning of MGA cDNA to allow expression analysis of hydrolytic and binding sites for this and other intestinal glucohydrolase enzyme activities. Methods: We sequenced 4 CNBr peptides and the N terminus from MGA isolated from human mucosa with HBB 2/143/17 mAb. RT used human intestinal RNA. PCR used degenerate primers with specific primers for extensions. Results: A 6515 bp cDNA coding a 1853 aa, 209 kDa protein was cloned. 92% identity was found between the sequence deduced from the cDNA and the purified peptide sequences. MGA peptide has 2 WIDMNE hydrolytic sites and the maltase and glucoamylase domains have many additional internal homologies. We are now expressing recombinant maltase and glucoamylase in E. coli and COS-1 cells in order to determine maltose and starch specificities of the isolated hydrolytic domains. Conclusions: MGA is a member of the Glycosyl Hydrolase Family 31. The MGA WIDMNE sequences, shared by sucrase-isomaltase, suggest a common mucosal hydrolytic mechanism with differences in the starch binding regions responsible for glucoamylase specificitiy.