Enzymatic Activity of the Staphylococcus aureus SplB Serine Protease is Induced by Substrates Containing the Sequence Trp-Glu-Leu-Gln

Dubin, Grzegorz; Steć-Niemczyk, Justyna; Kisielewska, Magdalena; Pustelny, Katarzyna; Popowicz, Grzegorz M.; Bista, Michal; Kantyka, Tomasz; Boulware, Kevin T.; Stennicke, Henning R.; Czarna, Anna; Phopaisarn, M.; Daugherty, Patrick S.; Thøgersen, Ida B.; Enghild, Jan J.; Thornberry, Nancy A.; Dubin, Adam; Potempa, Jan

doi:10.1016/j.jmb.2008.03.059

Cited by 43 publications

(69 citation statements)

References 68 publications

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“…More efficient substrates (Ac-WELD-ACC and Ac-WELQ-ACC) were later synthesized based on the consensus sequence recognized and cleaved by SplB as determined previously (13). Moreover, FRET protein substrates based on the SplB consensus sequence were used for assessing the kinetics of mutants.…”

Section: Methodsmentioning

confidence: 99%

“…Activity of SplB Protease Variants toward Consensus Sequence Substrates-The results described above were obtained using a peptide substrate (Ac-WELQ-ACC) based on the consensus sequence recognized and cleaved by SplB protease (WELQ2; 2 indicates the cleavage site) as established previously (13). To determine whether the differences in the activity of SplB protease variants constitute a general property of the enzyme, rather than being substrate-specific, we tested further two fluorescent peptide substrates and developed a protein-based FRET substrate.…”

Section: Figure 1 Removal Of Artificial N-terminal Extension Releasementioning

confidence: 99%

“…Crystal Structure Provides Insights into the Molecular Mechanism of Protease Activation-To reveal the molecular basis of the activating effect of N-terminal processing in GS-SplB, we determined the x-ray structure of the protein and compared it with a previously determined structure of SplB (PDB code 2VID) (13). Crystals of GS-SplB were obtained from two different conditions and belonged to different space groups.…”

Section: Figure 1 Removal Of Artificial N-terminal Extension Releasementioning

confidence: 99%

“…Moreover, the N-terminal amine of Glu-1 seems to transiently interact directly with loop 2 in SplB but not GS-SplB. Because loop 2 forms a part of the SplB protease active site being involved in substrate recognition (13), one may expect some influence on the process. However, because a large part of loop 2 is poorly defined by electron density and the entire region is rather flexible, this conclusion is highly ambiguous.…”

Section: Figure 1 Removal Of Artificial N-terminal Extension Releasementioning

confidence: 99%

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Staphylococcal SplB Serine Protease Utilizes a Novel Molecular Mechanism of Activation

Pustelny

Zdzalik

Stach

et al. 2014

Journal of Biological Chemistry

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Background: Activation of SplB protease requires precise N-terminal processing, but the molecular mechanism remains unknown. Results:The new N-terminal Glu-1 forms a distinctive H-bond network essential for full catalytic activity. Conclusion: Changes in protein dynamics rather than a direct effect on the active site are of crucial importance in SplB activation. Significance: A novel serine protease activation mechanism was uncovered.

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Section: Methodsmentioning

confidence: 99%

Section: Figure 1 Removal Of Artificial N-terminal Extension Releasementioning

confidence: 99%

Section: Figure 1 Removal Of Artificial N-terminal Extension Releasementioning

confidence: 99%

Section: Figure 1 Removal Of Artificial N-terminal Extension Releasementioning

confidence: 99%

See 2 more Smart Citations

Staphylococcal SplB Serine Protease Utilizes a Novel Molecular Mechanism of Activation

Pustelny

Zdzalik

Stach

et al. 2014

Journal of Biological Chemistry

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“…To date, SplA, SplB, SplC, and SplD are the best-characterized Spl proteases in terms of biochemical and structural properties and show significant structural homology to V8 protease and epidermolytic toxins (Popowicz et al, 2006; Dubin et al, 2008; Stec-Niemczyk et al, 2009; Zdzalik et al, 2013). By way of example, SplA shows a chymotripsin-like fold and consists of two domains, each of which is made up of six antiparallel β strands folded into a β barrel.…”

Section: S Aureus Expresses a Variety Of Proteases That Play A Role mentioning

confidence: 99%

Staphylococcus aureus Manipulates Innate Immunity through Own and Host-Expressed Proteases

Pietrocola

Nobile

Rindi

et al. 2017

Front. Cell. Infect. Microbiol.

122

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Neutrophils, complement system and skin collectively represent the main elements of the innate immune system, the first line of defense of the host against many common microorganisms. Bacterial pathogens have evolved strategies to counteract all these defense activities. Specifically, Staphylococcus aureus, a major human pathogen, secretes a variety of immune evasion molecules including proteases, which cleave components of the innate immune system or disrupt the integrity of extracellular matrix and intercellular connections of tissues. Additionally, S. aureus secretes proteins that can activate host zymogens which, in turn, target specific defense components. Secreted proteins can also inhibit the anti-bacterial function of neutrophils or complement system proteases, potentiating S. aureus chances of survival. Here, we review the current understanding of these proteases and modulators of host proteases in the functioning of innate immunity and describe the importance of these mechanisms in the pathology of staphylococcal diseases.

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Evolutionary optimization of peptide substrates for proteases that exhibit rapid hydrolysis kinetics

Boulware

Jabaiah

Daugherty

2010

Biotech & Bioengineering

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Protease cleavage site recognition motifs can be identified using protease substrate discovery methodologies, but typically exhibit non-optimal specificity and activity. To enable evolutionary optimization of substrate cleavage kinetics, a two-color cellular library of peptide substrates (CLiPS) methodology was developed. Two-color CLiPS was applied to identify peptide substrates for the tobacco etch virus (TEV) protease from a random pentapeptide library, which were then optimized by screening of a focused, extended substrate library. Quantitative library screening yielded seven amino acid substrates exhibiting rapid hydrolysis by TEV protease and high sequence similarity to the native seven-amino-acid substrate, with a strong consensus of EXLYPhiQG. Comparison of hydrolysis rates for a family of closely related substrates indicates that the native seven-residue TEV substrate co-evolved with TEV protease to facilitate highly efficient hydrolysis. Consensus motifs revealed by screening enabled database identification of a family of related, putative viral protease substrates. More generally, our results suggest that substrate evolution using CLiPS may be useful for optimizing substrate selectivity and activity to enable the design of more effective protease activity probes, molecular imaging agents, and prodrugs.

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Enzymatic Activity of the Staphylococcus aureus SplB Serine Protease is Induced by Substrates Containing the Sequence Trp-Glu-Leu-Gln

Cited by 43 publications

References 68 publications

Staphylococcal SplB Serine Protease Utilizes a Novel Molecular Mechanism of Activation

Staphylococcal SplB Serine Protease Utilizes a Novel Molecular Mechanism of Activation

Staphylococcus aureus Manipulates Innate Immunity through Own and Host-Expressed Proteases

Evolutionary optimization of peptide substrates for proteases that exhibit rapid hydrolysis kinetics

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