Generalities
The Aeromonas Sobria Serine Protease ASP protein is a serine protease that will cut peptide bonds after specific amino acids of a target protein. It preferentially cleaves peptide bonds that follow dibasic amino-acid residues. The kexin-like serine protease belongs to the subtilisin family (Subtilase). The structure of ASP is similar to that of Kex2 [1] (1r64), a protease of the subtilisin family, but ASP has a unique extra occluding region close to its active site.
This protein is secreted by the Anaerobic bacterium Aeromonas Sobria, which can cause potentially lethal septic shock. Septic Shock is a clinical syndrome of potentially fatal organ dysfunction caused by a disorder in the response to infection. In septic shock, there is a critical reduction in tissue perfusion; acute multivisceral failure, including the lungs, kidneys and liver, can be observed.
ASP is a sepsis-related factor. It can cause several dysfunction like by inducing vascular leakage, reducing blood pressure via the activation of the kinin system or promoting human plasma coagulation through the activation of prothrombin. Finally it can causes the formation of pus and edema through the action of anaphylatoxin C5a (4p3a). Gastroenteritis, and in extreme cases deuteropathy, are the main syndrome caused by infection with A.sobria.
The maturation of ASP is achieved by ORF2. This protein plays the role of an external chaperone and is necessary for the construction of the stable ASP. Indeed, ASP doesn’t contain a propeptide (such as Kex2) that is involved in the proper folding of the protein.
Phrase dans maturation à reformuler … “For maturation of ASP, the first 24 residues of the propeptide are cleaved and although a functional P-domain is reportedly necessary for maturation of the substitution domain in kexins”
Secondary structure
The structure of ASP is very similar to that of Kex2 (1r64), but it has a unique extra-occluding region close to its active site within the subtilisin domains. This extra-occluding region is unique and it could serve as a useful target to make the development of new antisepsis drugs easier.
The domain structure of ASP consists of the propeptide, the catalytic subtilisin-like domain, and the P-domain. The ASP molecule have two mean regions: an N-terminal region extending from Gly-3 to Pro-431 and forming the subtilisin domain, and a C-terminal region extending from Leu-432 to His-595 and forming the P-domain.
Moreover, we can find three Ca2+ Binding Sites in the ASP Structure (Ca1, Ca2 and Ca3). Ca1 and Ca2 are situated in the N-terminal domain, and Ca3 is situated in the C-terminal domain. It were assigned to ASP based on electron density, counter charges, and coordination. But in contrary to Kex2, ASP contains no Ca2 binding sites near its catalytic site. Those Ca2+ binding Site are important because ...
Domains
The Subtilisin Domain: It contains 10 helices (alpha 1 to 10) and twelve chains (béta 1 to 10 and béta 13 to 14). The N-terminal domain of ASP seems to be like the catalytic domain of Kex2, which is similar to those of subtilisin and other subtilisin-related proteases. This ASP catalytic site contains the catalytic Asp-78, His-115, and Ser-336 residues characteristic of subtilisins. In addition, 4 loops (L) protrude from the N-terminal subtilisin domain of ASP: Gly-3– Pro-26 (L1), Asn-221–Phe-241 (L2), Gly-300–Cys-326 (L3), and Gln-377–Glu-397 (L4). L1, L2, and L3 have random coil structure, whereas L4 forms a hairpin that protrudes toward the P-domain. Moreover, two disulfide bridges are formed between Cys-4 and Cys-24 in L1 and between Cys-301 and Cys-326 in L3, which stabilize those loops.
The P-domain: The core of the P-domain in ASP contains 8 béta-strands (béta 16 18 23 and 26). The extra occluding-region is comprised of two parts, pL1(Gly 521–Thr 525, béta 5, 6, and 12) and pL2 (Gly-557–Asn-578, béta 25), and it is situated close to the catalytic triad Asp-78,His-115,and Ser-336.
Active site
The catalytic triad: The catalytic triad of ASP is composed of Asp78, His115 and Ser336. These amino acids are the base is the active site of the protein, where the mode of action of the serine protease takes place.
A peptide can be inserted in the space of the active site. There, the amino acids of the catalytic triad will interact together and the mechanism will lead to a cut in the polypeptide.
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Mechanism: The mechanism is the following: The histidine will react with the serine and deprotonate it. The deprotonated hydroxyl group of the serine will act as a nucleophilic species and attack the carbon from the carbonyl function on the peptide. This will lead to the formation of a tetrahedral intermediate. Then, a second tetrahedral intermediate will be formed, but with the attack of a deprotonated water molecule. At the end, the regeneration of the active site will be done with the release of the peptide cut in two parts.
Disease
