Structural basis underlying the synergism of NADase and SLO during group A Streptococcus infection

Abstract

Group A Streptococcus (GAS) is a strict human pathogen possessing a unique pathogenic trait that utilizes the cooperative activity of NAD+-glycohydrolase (NADase) and Streptolysin O (SLO) to enhance its virulence. How NADase interacts with SLO to synergistically promote GAS cytotoxicity and intracellular survival is a long-standing question. Here, the structure and dynamic nature of the NADase/SLO complex are elucidated by X-ray crystallography and small-angle scattering, illustrating atomic details of the complex interface and functionally relevant conformations. Structure-guided studies reveal a salt-bridge interaction between NADase and SLO is important to cytotoxicity and resistance to phagocytic killing during GAS infection. Furthermore, the biological significance of the NADase/SLO complex in GAS virulence is demonstrated in a murine infection model. Overall, this work delivers the structure-functional relationship of the NADase/SLO complex and pinpoints the key interacting residues that are central to the coordinated actions of NADase and SLO in the pathogenesis of GAS infection.

Streptolysin O Accelerates the Conversion of Plasminogen to Plasmin

“…Similar movement of this domain has previously been reported for SLO-NADase complex. 52 Given that a crystal structure may not fully represent the dynamic nature of protein movement in solution, the reported crosslinks are likely derived from an ensemble of different conformations, including both bound and unbound states. Reciprocally, the overlength cross-links within PLG indicate local movement in particularly PAN, K4 and PSD domains.…”

PRMT-7/PRMT7 activates HLH-30/TFEB to guard plasma membrane integrity compromised by bacterial pore-forming toxins

“…Caco-2 and HeLa cells were incubated with DMEM containing 0.4 μg/mL SLO, prepared as described [ 53 ], and 5 mM dithiothreitol (DTT) (Sigma-Aldrich, 3 December 3483), for the reduction of disulfide bonds in streptolysin O (SLO), on ice for 30 min. Low temperatures promote SLO and cholesterol binding on the cell surface.…”

RopB-regulated SpeB cysteine protease degrades extracellular vesicles-associated streptolysin O and bacterial proteins from group A Streptococcus

“… Plasmid or strain Description a Reference or source Plasmids pCN143 Temperature-sensitive, E . coli -streptococci shuttle vector [ 42 ] pCN144 pCN143: speB T574A [ 43 ] pCN171 pCN143: slo ∆ cat [ 44 ] pCN226 pCN143: speB ∆ cat This study Strains A20 The emm 1 wild-type strain [ 41 ] SCN142 (∆ ropB ) A20 ropB isogenic mutant [ 13 ] SCN138 (SpeB C192S ) A20 SpeB C192S substituted mutant [ 43 ] SCN305 ( SIP *) A20 SIP-inactivated mutant [ 12 ] SCN344 (∆ speB ) A20 speB isogenic mutant This study SCN356 (∆ slo ) A20 slo isogenic mutant [ 44 ] Note: a , cat , chloramphenicol cassette. …”

“…The speB gene in this plasmid was removed by PCR with the reverse primers a-speB-SacII-F (5’-tagccgcggtatggaaatgcatttcg-3’) and a-speB-SacII-R (5’-catccgcggtttttttatacctcttt-3’) and replaced by the chloramphenicol cassette from Vector-78 [ 48 ]. The constructed plasmid, pCN226, and pCN171 (for construction of the slo mutant) [ 44 ] and pCN144 (for construction of SpeB C192S substitution) [ 45 ] were transformed into the wild-type A20 strain by electroporation and the transformants were selected as described previously [ 42 ]. The deletions and mutations in the target genes were confirmed by Sanger sequencing.…”