Heat labile enterotoxin from enterotoxigenic Escherichia coli is similar to cholera toxin (CT) and is a leading cause of diarrhea in developing countries. It consists of an enzymatically active A subunit (LTA) and a carrier pentameric B subunit (LTB). In the current study, we evaluated the importance of the N-terminal region of LTB by mutation analysis. Deletion of the glutamine (Delta Q3) residue and a substitution mutation E7G in the alpha 1 helix region led to defects in LTB protein secretion. Deletion of the proline residue (Delta P2) caused a decrease in alpha helicity. The Delta P2 mutant affected GM, ganglioside receptor binding activity without affecting LTB pentamer formation. Upon refolding/reassembly, the Delta P2 mutant showed defective biological activity. The single substitution mutation (E7D) strengthened the helix, imparting structural stability and thereby improved the GM] ganglioside receptor binding activity. Our results demonstrate the important role of N-terminal alpha 1 helix in maintaining the structural stability and the integrity of GM(1) ganglioside receptor binding activity. (C) 2008 Elsevier Inc. All rights reserved.