Aureolysin

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EC no. 3.4.24.29
CAS no. 39335-13-2
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Aureolysin (EC 3.4.24.29, protease III, staphylococcal metalloprotease,Staphylococcus aureus neutral proteinase) is an extracellular metalloprotease expressed by Staphylococcus aureus . [1] [2] [3] [4] [5] This protease is a major contributor to the bacterium's virulence, or ability to cause disease, by cleaving host factors of the innate immune system as well as regulating S. aureus secreted toxins and cell wall proteins. [6] [7] To catalyze its enzymatic activities, aureolysin requires zinc and calcium which it obtains from the extracellular environment within the host. [6] [7]

Contents

Genetics

Aureolysin is expressed from the gene aur, which is located on a monocistronic operon. [8] The gene exists in two allelic forms but the sequence is highly conserved with 89% homology between the two. [9] The gene contains a coding sequence of 1,527 nucleotides that translates into a pre-pro-form of the enzyme that is 509 amino acids long. [9] Of the 509 amino acids, only 301 denote the mature form of aureolysin. [9] After translation, the pre-portion of the enzyme is a 27 amino acid N-terminal signal peptide that acts as a guide to the secretion system located within the cell wall. [9] Here, the signal peptide is cleaved upon secretion of aureolysin. [9]

Aureolysin is largely co-expressed with other major proteases of S. aureus including the two cystine proteases, Staphopain A (ScpA) and B (SspB), and a serine protease V8 (SspA). The transcriptional regulation of aur is controlled by "housekeeping"  sigma factor  σA, and is up-regulated by accessory gene regulator agr. Expression levels of aureolysin are at their highest during the post-exponential phase however, up-regulation of aureolysin during phagocytosis has also been observed. [10] Transcription is repressed by staphylococcal accessory regulator sarA and by alternative sigma factor σB (a stress response modulator of  Gram-positive bacteria). 

The aur gene has a high prevalence in the genome of both  commensal- and  pathogenic-type S. aureus  strains. [11]

Activation

The staphylococcal proteolytic cascade. Once aureolysin, V8, and SspB are secreted into the extracellular environment, the pro-peptides, as indicated by the stripped smaller circles, need to be cleaved to generate enzyme activity. Aureolysin undergoes autocatalysis to remove the pro-peptide then cleaves V8, generating mature protease. V8 then activates SspB and the cascade is complete. Staphylococcal Proteolytic Cascade.jpg
The staphylococcal proteolytic cascade. Once aureolysin, V8, and SspB are secreted into the extracellular environment, the pro-peptides, as indicated by the stripped smaller circles, need to be cleaved to generate enzyme activity. Aureolysin undergoes autocatalysis to remove the pro-peptide then cleaves V8, generating mature protease. V8 then activates SspB and the cascade is complete.

Aureolysin, along with V8, SspB, and ScpA, are all secreted a zymogens. This means that they are secreted in an inactive conformation until the propeptide is removed in some manner. Aureolysin, V8, and SspB constitute what is known as the staphylococcal proteolytic cascade. [8] All three of these proteases are secreted into the environment with the propeptide inhibiting their activation. Aureolysin undergoes autocatalysis and the propeptide is degraded generating the mature form of the enzyme. [8] Mature aureolysin will then cleave the propeptide from V8, causing this protease to become active. [8] Finally V8 will cleave SspB propeptide and the cascade is now complete. [8] ScpA becomes mature by autocatalytic degradation of the propeptide, similar to that of aureolysin. [8]

The active residues of aureolysin are of critical importance to its enzymatic function. [9] The active residue is a glutamate amino acid located at the 145th position of the protein. [9]

Immune Evasion

Aureolysin cleaves various immune components and host proteins. It is important for hiding the bacterium from the immune system and is responsible for mediating the transition of a biofilm forming phenotype to a mobile and invasive one. There are many different targets of aureolysin and the effect on each is critical for the bacterium's virulence.

One major way aureolysin contributes to infection, is by inactivating certain targets within the complement system. Of all the proteases, aureolysin is the most effective against the complement cascade. [12] In all three pathways of complement activation, there is a target for the protease to manipulate. In the classical pathway, aureolysin not only decreases deposition of C1q on the S. aureus bacterial surface, it induces C1q to bind surfaces and deposit on commensal bacteria surfaces that typically do not activate the innate immune system. [12] Aureolysin has also been noted to produce high levels of C5a in human plasma, which leads to overstimulation of neutrophils that ultimately results in neutrophil death. [12] C3 is another major target of aureolysin. The active site has a high affinity for C3 and will cleave it into C3a and C3b however, the protein is cleaved two amino acid residues away from the native site that is recognized by the host C3 convertase. [7] [12] The aureolysin derived C3a and C3b are further degraded by host complement inhibitor factor H and I. [7] [12] In the lectin pathway, aureolysin inhibits MBL and ficolin binding which, in turn, reduce C3b deposition. [6]

Further immune evasion outside of the complement system occurs in various ways. Aureolysin cleaves and inactivates protease inhibitor α1-antichymotrypsin and partially inactivates α1-antitrypsin. [13] The cleavage of α1-antitrypsin generates a fragment chemotactic to neutrophils, and the cleavage of both protease inhibitors causes deregulation of neutrophil-derived proteolytic activity. [13] Aureolysin has also been shown to cleave the antimicrobial peptide LL-37, rendering it inactive and unable to puncture the bacterial cell wall. Production of immunoglobulin by lymphocytes is inhibited by aureolysin as well. [5] It contributes to both coagulation triggered by coagulase, and to fibrinolysis mediated by staphylokinase. [13] Proteolytic conversion of pro-thrombin into thrombin by aureolysin works synergistically with coagulase and contributes to the staphylocoagulation of human plasma. [13] By inducing staphylocoagulation, the bacterium is hidden within the clot from phagocytic cells. Contradictory to staphylocoagulation, aureolysin is responsible for the activation of urokinase, and inactivation of α2-antiplasmin and plasminogen activator inhibitor-1. [7] This promotes the dissemination of the bacterium to allow for further invasion of the host.

Biological significance

When S. aureus is establishing an infection within a host, it needs to continuously switch from a static, or biofilm forming phenotype, to an invasive, or mobile phenotype. The proteases help mediate this process. Aureolysin appears to down-regulate the formation of biofilms and allow for the mobility of the bacterium. One way it contributes to this change is by mediating coagulation as well as the activation of urokinase. However, it also mediates S. aureus cell wall and secreted proteins to promote this change. For example, clumping factor B is a surface protein that is responsible for the binding of fibrinogen around the bacterium to hide it within a clot. [5] [11] Aureolysin is responsible for the cleavage of clumping factor B, which causes the loss of S. aureus binding to fibrinogen. By this mechanism, it may act as a self-regulatory mechanism for dissemination and spreading in combination with activation of fibrinolysis, while the protease simultaneously provides protection against complement activation. [5] [11] It has been demonstrated that aureolysin has impact for bacterial survival in human whole blood. [14] Aureolysin is also up-regulated upon phagocytosis and promotes intracellular survival. [5] [10] [15]

S. aureus prefers to establish a chronic, or long lasting infection within a host. While promoting dissemination and counteracting immune mechanisms, aureolysin also regulates secreted virulence factors to control the pathogenicity of the bacterium. By inactivation of PSMs and α-toxins, aureolysin may suppress the pathogenic impact of the bacteria allowing for a chronic infection to be established. [5]

Related Research Articles

<span class="mw-page-title-main">Proteolysis</span> Breakdown of proteins into smaller polypeptides or amino acids

Proteolysis is the breakdown of proteins into smaller polypeptides or amino acids. Uncatalysed, the hydrolysis of peptide bonds is extremely slow, taking hundreds of years. Proteolysis is typically catalysed by cellular enzymes called proteases, but may also occur by intra-molecular digestion.

<span class="mw-page-title-main">Protease</span> Enzyme that cleaves other proteins into smaller peptides

A protease is an enzyme that catalyzes proteolysis, breaking down proteins into smaller polypeptides or single amino acids, and spurring the formation of new protein products. They do this by cleaving the peptide bonds within proteins by hydrolysis, a reaction where water breaks bonds. Proteases are involved in numerous biological pathways, including digestion of ingested proteins, protein catabolism, and cell signaling.

<i>Staphylococcus aureus</i> Species of Gram-positive bacterium

Staphylococcus aureus is a Gram-positive spherically shaped bacterium, a member of the Bacillota, and is a usual member of the microbiota of the body, frequently found in the upper respiratory tract and on the skin. It is often positive for catalase and nitrate reduction and is a facultative anaerobe that can grow without the need for oxygen. Although S. aureus usually acts as a commensal of the human microbiota, it can also become an opportunistic pathogen, being a common cause of skin infections including abscesses, respiratory infections such as sinusitis, and food poisoning. Pathogenic strains often promote infections by producing virulence factors such as potent protein toxins, and the expression of a cell-surface protein that binds and inactivates antibodies. S. aureus is one of the leading pathogens for deaths associated with antimicrobial resistance and the emergence of antibiotic-resistant strains, such as methicillin-resistant S. aureus (MRSA), is a worldwide problem in clinical medicine. Despite much research and development, no vaccine for S. aureus has been approved.

<span class="mw-page-title-main">Phagocyte</span> Cells that ingest harmful matter within the body

Phagocytes are cells that protect the body by ingesting harmful foreign particles, bacteria, and dead or dying cells. Their name comes from the Greek phagein, "to eat" or "devour", and "-cyte", the suffix in biology denoting "cell", from the Greek kutos, "hollow vessel". They are essential for fighting infections and for subsequent immunity. Phagocytes are important throughout the animal kingdom and are highly developed within vertebrates. One litre of human blood contains about six billion phagocytes. They were discovered in 1882 by Ilya Ilyich Mechnikov while he was studying starfish larvae. Mechnikov was awarded the 1908 Nobel Prize in Physiology or Medicine for his discovery. Phagocytes occur in many species; some amoebae behave like macrophage phagocytes, which suggests that phagocytes appeared early in the evolution of life.

<span class="mw-page-title-main">Classical complement pathway</span> Aspect of the immune system

The classical complement pathway is one of three pathways which activate the complement system, which is part of the immune system. The classical complement pathway is initiated by antigen-antibody complexes with the antibody isotypes IgG and IgM.

Granzymes are serine proteases released by cytoplasmic granules within cytotoxic T cells and natural killer (NK) cells. They induce programmed cell death (apoptosis) in the target cell, thus eliminating cells that have become cancerous or are infected with viruses or bacteria. Granzymes also kill bacteria and inhibit viral replication. In NK cells and T cells, granzymes are packaged in cytotoxic granules along with perforin. Granzymes can also be detected in the rough endoplasmic reticulum, golgi complex, and the trans-golgi reticulum. The contents of the cytotoxic granules function to permit entry of the granzymes into the target cell cytosol. The granules are released into an immune synapse formed with a target cell, where perforin mediates the delivery of the granzymes into endosomes in the target cell, and finally into the target cell cytosol. Granzymes are part of the serine esterase family. They are closely related to other immune serine proteases expressed by innate immune cells, such as neutrophil elastase and cathepsin G.

<span class="mw-page-title-main">Phagosome</span>

In cell biology, a phagosome is a vesicle formed around a particle engulfed by a phagocyte via phagocytosis. Professional phagocytes include macrophages, neutrophils, and dendritic cells (DCs).

<span class="mw-page-title-main">Innate immune system</span> One of the two main immunity strategies

The innate, or nonspecific, immune system is one of the two main immunity strategies in vertebrates. The innate immune system is an alternate defense strategy and is the dominant immune system response found in plants, fungi, insects, and primitive multicellular organisms.

<span class="mw-page-title-main">Complement component 5a</span> Protein fragment

C5a is a protein fragment released from cleavage of complement component C5 by protease C5-convertase into C5a and C5b fragments. C5b is important in late events of the complement cascade, an orderly series of reactions which coordinates several basic defense mechanisms, including formation of the membrane attack complex (MAC), one of the most basic weapons of the innate immune system, formed as an automatic response to intrusions from foreign particles and microbial invaders. It essentially pokes microscopic pinholes in these foreign objects, causing loss of water and sometimes death. C5a, the other cleavage product of C5, acts as a highly inflammatory peptide, encouraging complement activation, formation of the MAC, attraction of innate immune cells, and histamine release involved in allergic responses. The origin of C5 is in the hepatocyte, but its synthesis can also be found in macrophages, where it may cause local increase of C5a. C5a is a chemotactic agent and an anaphylatoxin; it is essential in the innate immunity but it is also linked with the adaptive immunity. The increased production of C5a is connected with a number of inflammatory diseases.

Virulence factors are cellular structures, molecules and regulatory systems that enable microbial pathogens to achieve the following:

<span class="mw-page-title-main">Hemolysin</span> Molecule destroying the membrane of red blood cells

Hemolysins or haemolysins are lipids and proteins that cause lysis of red blood cells by disrupting the cell membrane. Although the lytic activity of some microbe-derived hemolysins on red blood cells may be of great importance for nutrient acquisition, many hemolysins produced by pathogens do not cause significant destruction of red blood cells during infection. However, hemolysins are often capable of lysing red blood cells in vitro.

Porphyromonas gingivalis belongs to the phylum Bacteroidota and is a nonmotile, Gram-negative, rod-shaped, anaerobic, pathogenic bacterium. It forms black colonies on blood agar.

<span class="mw-page-title-main">ADAMTS3</span> Protein-coding gene in the species Homo sapiens

A disintegrin and metalloproteinase with thrombospondin motifs 3 is an enzyme that in humans is encoded by the ADAMTS3 gene. The protein encoded by this gene is the major procollagen II N-propeptidase.

'Staphylococcus aureus delta toxin is a toxin produced by Staphylococcus aureus. It has a wide spectrum of cytolytic activity.

Omptins are a family of bacterial proteases. They are aspartate proteases, which cleave peptides with the use of a water molecule. Found in the outer membrane of gram-negative enterobacteria such as Shigella flexneri, Yersinia pestis, Escherichia coli, and Salmonella enterica. Omptins consist of a widely conserved beta barrel spanning the membrane with 5 extracellular loops. These loops are responsible for the various substrate specificities. These proteases rely upon binding of lipopolysaccharide for activity.

<span class="mw-page-title-main">OmpT</span> Bacterial protein

OmpT is an aspartyl protease found on the outer membrane of Escherichia coli. OmpT is a subtype of the family of omptin proteases, which are found on some gram-negative species of bacteria.

<span class="mw-page-title-main">Glutamyl endopeptidase GluV8</span>

Glutamyl endopeptidase is an extracellular bacterial serine protease of the glutamyl endopeptidase I family that was initially isolated from the Staphylococcus aureus strain V8. The protease is, hence, commonly referred to as "V8 protease", or alternatively SspA from its corresponding gene.

Plasminogen activator Pla is an enzyme. This enzyme catalyses the following chemical reaction

Glutamyl endopeptidase I is a family of extracellular bacterial serine proteases. The proteases within this family have been identified in species of Staphylococcus, Bacillus, and Streptomyces, among others. The two former are more closely related, while the Streptomyces-type is treated as a separate family, glutamyl endopeptidase II.

Staphopain A (<i>Staphylococcus aureus</i>)

Staphopain A is a secreted cysteine protease produced by Staphylococcus aureus. It was first identified in the S. aureus V8 strain as a papain-like cysteine protease. The protease distinguishes itself from the other major proteases of S. aureus in its very broad specificity and its ability to degrade elastin.

References

  1. ↑ Arvidson, S. (1973). "Studies on extracellular proteolytic enzymes from Staphylococcus aureus. II. Isolation and characterization of an EDTA-sensitive protease". Biochim. Biophys. Acta. 302 (1): 149–157. doi:10.1016/0005-2744(73)90017-x. PMID   4632563.
  2. ↑ Saheb SA (1976). "Purification and characterization of an extracellular protease from Staphylococcus aureus inhibited by EDTA". Biochimie (in French). 58 (7): 793–804. doi:10.1016/s0300-9084(76)80310-0. PMID   823980.
  3. ↑ Drapeau GR (November 1978). "Role of metalloprotease in activation of the precursor of staphylococcal protease". Journal of Bacteriology. 136 (2): 607–13. doi:10.1128/JB.136.2.607-613.1978. PMC   218585 . PMID   711676.
  4. ↑ Potempa J, Porwit-Bobr Z, Travis J (December 1989). "Stabilization vs. degradation of Staphylococcus aureus metalloproteinase". Biochimica et Biophysica Acta (BBA) - General Subjects. 993 (2–3): 301–4. doi:10.1016/0304-4165(89)90181-5. PMID   2512988.
  5. 1 2 3 4 5 6 Potempa J, Shaw LN (2013-01-01). Rawlings ND, Salvesen G (eds.). Handbook of Proteolytic Enzymes. Academic Press. pp. 563–569. doi:10.1016/b978-0-12-382219-2.00114-9. ISBN   9780123822192.
  6. 1 2 3 Laarman, Alexander J.; et al. (April 2011). "Staphylococcus aureus Metalloprotease Aureolysin Cleaves Complement C3 To Mediate Immune Evasion". Journal of Immunology. 186 (11): 6445–6453. doi: 10.4049/jimmunol.1002948 . PMID   21502375.
  7. 1 2 3 4 5 Pietrocola, Giampiero; et al. (30 August 2017). "Staphylococcus aureus Manipulates Innate Immunity through Own and Host-Expressed Proteases". Frontiers in Cellular and Infection Microbiology. 7: 166. doi: 10.3389/fcimb.2017.00166 . PMC   5418230 . PMID   28529927.
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  9. 1 2 3 4 5 6 7 Sabat, A; et al. (February 2000). "Two Allelic Forms of Aureolysin Gene (aur) within Staphylococcus aureus". Infection and Immunity. 68 (2): 973–976. doi:10.1128/IAI.68.2.973-976.2000. PMC   97234 . PMID   10639475.
  10. 1 2 Burlak C, Hammer CH, Robinson MA, Whitney AR, McGavin MJ, Kreiswirth BN, Deleo FR (May 2007). "Global analysis of community-associated methicillin-resistant Staphylococcus aureus exoproteins reveals molecules produced in vitro and during infection". Cellular Microbiology. 9 (5): 1172–90. doi:10.1111/j.1462-5822.2006.00858.x. PMC   2064037 . PMID   17217429.
  11. 1 2 3 Dubin G (2002-07-01). "Extracellular proteases of Staphylococcus spp". Biological Chemistry. 383 (7–8): 1075–86. doi:10.1515/BC.2002.116. PMID   12437090. S2CID   23295763.
  12. 1 2 3 4 5 Jusko, Monika; Potempa, Jan; Kantyka, Tomasz; Bielecka, Ewa; Miller, Halie K.; Kalinska, Magdalena; Dubin, Grzegorz; Garred, Peter; Shaw, Lindsey N. (2014). "Staphylococcal Proteases Aid in Evasion of the Human Complement System". Journal of Innate Immunity. 6 (1): 31–46. doi:10.1159/000351458. ISSN   1662-8128. PMC   3972074 . PMID   23838186.
  13. 1 2 3 4 Dubin, Grzegorz (July 2002). "Extracellular proteases of Staphylococcus spp". Biological Chemistry. 383 (7–8): 1075–1086. doi:10.1515/BC.2002.116. ISSN   1431-6730. PMID   12437090. S2CID   23295763.
  14. ↑ Jusko M, Potempa J, Kantyka T, Bielecka E, Miller HK, Kalinska M, Dubin G, Garred P, Shaw LN, Blom AM (2014-01-01). "Staphylococcal proteases aid in evasion of the human complement system". Journal of Innate Immunity. 6 (1): 31–46. doi:10.1159/000351458. PMC   3972074 . PMID   23838186.
  15. ↑ Kubica M, Guzik K, Koziel J, Zarebski M, Richter W, Gajkowska B, Golda A, Maciag-Gudowska A, Brix K, Shaw L, Foster T, Potempa J (January 2008). "A potential new pathway for Staphylococcus aureus dissemination: the silent survival of S. aureus phagocytosed by human monocyte-derived macrophages". PLOS ONE. 3 (1): e1409. Bibcode:2008PLoSO...3.1409K. doi: 10.1371/journal.pone.0001409 . PMC   2169301 . PMID   18183290.
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