The autotransporter mode of surface presentation in Gram-negative bacteria requires a hypothetical C-terminal P-barrel which makes up an aqueous channel in the outer membrane. PaIA is a Pseudomonas sp. autotransporter lipolytic protein. PaIA is a 66 kDa protein that is composed of two parts, the N-terminal region (Ala(1)-Ala(296)) similar to the GDSL lipases and the C-terminal region (Leu(320)-Phe(612)) to the autotransporter. In this report, we provide biochemical and structural evidence demonstrating that the pore size of the P-barrel conduit is important in delivering the N-terminal domain to the cell surface. Among all the autotransporter domains two strictly conserved residues (Pro(478) and Gly(576) in PaIA) are converted to other various residues using site-directed mutagenesis. This investigation was made into the different pore-size mutants, affecting the folding of N-terminal domain. Wild P-domain contains a cavity of similar to2 mn diameter that is optimal for the active conformation of the N-terminal domains. However, deviation from the proper size of the pore, whether it is larger or smaller, is not suitable for the proper folding of the N-terminal catalytic domain. (C) 2003 Elsevier Inc. All rights reserved.