Characterization of Streptococcus gordonii SecA2 as a Paralogue of SecA

Abstract

The accessory Sec system of Streptococcus gordonii is essential for transport of the glycoprotein GspB to the bacterial cell surface. A key component of this dedicated transport system is SecA2. The SecA2 proteins of streptococci and staphylococci are paralogues of SecA and are presumed to have an analogous role in protein transport, but they may be specifically adapted for the transport of large, serine-rich glycoproteins. We used a combination of genetic and biochemical methods to assess whether the S. gordonii SecA2 functions similarly to SecA. Although mutational analyses demonstrated that conserved amino acids are essential for the function of SecA2, replacing such residues in one of two nucleotide binding folds had only minor effects on SecA2 function. SecA2-mediated transport is highly sensitive to azide, as is SecA-mediated transport. Comparison of the S. gordonii SecA and SecA2 proteins in vitro revealed that SecA2 can hydrolyze ATP at a rate similar to that of SecA and is comparably sensitive to azide but that the biochemical properties of these enzymes are subtly different. That is, SecA2 has a lower solubility in aqueous solutions and requires higher Mg 2؉ concentrations for maximal activity. In spite of the high degree of similarity between the S. gordonii paralogues, analysis of SecA-SecA2 chimeras indicates that the domains are not readily interchangeable. This suggests that specific, unique contacts between SecA2 and other components of the accessory Sec system may preclude cross-functioning with the canonical Sec system.

Canonical SecA Associates with an Accessory Secretory Protein Complex Involved in Biogenesis of a Streptococcal Serine-Rich Repeat Glycoprotein

“…Biochemical analysis demonstrates that SecA2 is an ATPase that shares several structural features with SecA (2), suggesting that SecA2 may have a similar role in export of SRRP preproteins. Mutagenesis of SecA2 not only abolished the export of Fap1, GspB, and other SRRPs (1,7,19,24) but also inhibited secretion of a major virulent lipoprotein, FimA, of S. parasanguinis (4,7).…”

Asp2 and Asp3 Interact Directly with GspB, the Export Substrate of the Streptococcus gordonii Accessory Sec System

“…SecA is an ATPase motor protein that powers the transport of substrates through the membrane channel (translocon) formed by SecY in complex with SecE and SecG (14). SecA2 is also an ATPase and is likely to play a similar role in the transport of the accessory Sec substrate (7,13). Although SecY2 has not been studied extensively, it is predicted to share structural similarities to SecY and is likely to function analogous to SecY (6,8,42).…”

A prl Mutation in SecY Suppresses Secretion and Virulence Defects of Listeria monocytogenes secA2 Mutants

“…There are two types of SecA2 secretion pathways; some bacteria have an additional SecY homolog, which functions together with SecA2 to form a SecA2-SecY2 system. This system has been extensively studied in streptococci and is highly specialized to export glycosylated proteins that are incompatible with the canonical Sec system (13,(17)(18)(19)(20). Other bacteria contain a SecA2-only system, which is thought to interact with the canonical SecYEG channel (16,21,22).…”