LIPASE SECRETION BY BACTERIAL HYBRID ATP-BINDING CASSETTE EXPORTERS -MOLECULAR RECOGNITION OF THE LIPBCD, PRTDEF, AND HASDEF EXPORTERS

Serrartia marcescens secretes several proteins, such as the lipase Lip A, the metalloprotease PrtA, and the heme-binding protein HasA, which is required for heme acquisition, through two N-terminal signal peptid e independent systems that are classified as bacterial ATP-binding cas sette (ABC) exporters, One is the ABC exporter for HasA, consisting of the ABC protein HasD, the membrane fusion protein (MFP) HasE, and the outer membrane protein (OMP) HasF, The second, composed of LipB (an A BC protein), LipC (an MFP), and LipD (an OMP), promotes secretion of L ipA and PrtA in Escherichia coli recombinant clones, PrtA, which shows homology to the Erwinia chrysanthemi metalloproteases, is efficiently secreted by E. coli cells carrying the E. chrysanthemi ABC exporter P rtD (ABC protein)-PrtE (MFP)-PrtF (OMP), The existence of distinct sys tems in this bacterium and of various substrates for these systems all owed the study of protein secretion by heterologous Has, Lip, and Prt systems and by Has-Lip and Lip-Prt hybrid exporters in the genuine hos t as well as in E. coli, For that purpose, lipB-, lipC-, and lipD-defi cient mutants were isolated from S. marcescens 8000 and their secretio n of LipA and PrtA was analyzed. This demonstrated that a unique expor ter, the Lip apparatus, in S. marcescens secretes both LipA and PrtA. Hybrid exporters were tested for secretion of HasA and LipA, The LipB- HasE-HasF exporter allowed secretion of LipA but not HasA, showing tha t the ABC protein LipB is responsible for the substrate specificity, L ipA, HasA, and E. chrysanthemi PrtC were secreted via heterologous exp orters and via some hybrid exporters, Analysis of secretion via hybrid exporters showed that specific interactions occur between MFPs and OM Ps in these systems, These genetic experiments demonstrated that speci fic interactions between the ABC protein and the MFP are required for the formation of active exporters.