Abstract
Bacteroides thetaiotaomicron (Bt) is a prominent member of the human gut microbiota with an extensive capacity for glycan harvest. This bacterium expresses a five-protein complex in the outer membrane, called the starch utilization system (Sus), which binds, degrades, and imports starch into the cell. Sus is a model system for the many glycan-targeting polysaccharide utilization loci found in Bt and other members of the Bacteroidetes phylum. Our previous work has shown that SusG, a lipidated amylase in the outer membrane, explores the entire cell surface but diffuses more slowly as it interacts with starch. Here, we use a combination of single-molecule tracking, super-resolution imaging, reverse genetics, and proteomics to show that SusE and SusF, two proteins that bind starch, are immobile on the cell surface even when other members of the system are knocked out and under multiple different growth conditions. This observation suggests a new paradigm for protein complex formation: binding proteins form immobile complexes that transiently associate with a mobile enzyme partner.