The assembly landscape of the complete B-repeat superdomain fromStaphylococcus epidermidisstrain 1457

Abstract

AbstractThe accumulation-associated protein (Aap) is the primary determinant ofStaphylococcus epidermidisdevice-related infections. The B-repeat superdomain is responsible for intercellular adhesion that leads to the development of biofilms occurring in such infections. It was recently demonstrated that Zn-induced B-repeat assembly leads to formation of functional amyloid fibrils, which offer strength and stability to the biofilm. Rigorous biophysical studies of Aap B-repeats fromS. epidermidisstrain RP62A revealed Zn-induced assembly into a stable, reversible dimer and tetramer, prior to aggregation into amyloid fibrils. Genetic manipulation is not tractable for manyS. epidermidisstrains, including RP62A; instead, many genetic studies have used strain 1457. Therefore, to better connect findings from biophysical and structural studies of B-repeats toin vivostudies, the B-repeat superdomain from strain 1457 was examined. Differences between the B-repeats from strain RP62A and 1457 include the number of B-repeats, which has been shown to play a critical role in assembly into amyloid fibrils, as well as the distribution of consensus and variant B-repeat subtypes, which differ in assembly competency and thermal stability. Detailed investigation of the Zn-induced assembly of the full B-repeat supderdomain from strain 1457 was conducted using analytical ultracentrifugation. Whereas the previous construct from RP62A (Brpt5.5) formed a stable tetramer prior to aggregation, Brpt6.5 from 1457 forms extremely large stable species on the order of 28- and 32-mers, prior to aggregation into similar amyloid fibrils. Importantly, both assembly pathways proceed through the same mechanism of dimerization and tetramerization, and both conclude with the formation of amyloid-like fibrils. Theoretical discussions of the energy landscape of B-repeats from different strains and of different length are provided with considerations of biological implications.Statement of SignificanceStaphylococcus epidermidisis a major pathogen responsible for device-related infections. The primary factor responsible for such infections is the accumulation-associated protein, Aap, through its ability to mediate formation of dense surface-adherent communities of bacteria known as biofilms. Our lab recently demonstrated that the B-repeat superdomain of Aap from strain RP62A undergoes Zn-dependent assembly to form functional amyloid fibrils that improve the strength and resilience of biofilmsin vitro. These amyloid fibrils may be responsible for the difficulty in treating device-related infections. However, strain 1457 is commonly used for genetic manipulation. In this manuscript, the Zn-dependent assembly of B-repeats from strain 1457 is shown to lead to the same outcome of amyloidogenesis, although it occurs through different intermediate oligomeric states.