Structural basis of the autolysis of AaHIV suggests a novel target recognizing model for ADAM/reprolysin family proteins

AaHIV, a P-III-type snake venom metalloproteinase (SVMP), consists of metalloproteinase/disintegrin/cysteine-rich (MDC) domains and is homologous to a disintegrin and metalloproteinase (ADAM) family proteins. Similar to brevilysin H6 and jararhagin, AaHIV can easily autolyse to release a stable protein named acucetin, which contains disintegrin-like and cysteine-rich domains. In this study, we determined the crystal structure of AaHIV and investigated the autolysis mechanism. Based on the structure of AaHIV and the results from docking experiments, we present a new model for target recognition in which two protein molecules form a functional unit, and the DC domain of one molecule is used for target recognition while the M-domain of the other is used for target proteolysis. Our results shed new light on the mechanism of target recognition and processing in ADAM/reprolysin family proteins.     

Autolysis at the disintegrin domain of patagonfibrase,a metalloproteinase from Philodryas patagoniensis (Patagonia Green Racer; Dipsadidae) venom

Patagonfibrase is a 57.5-kDa hemorrhagic metalloproteinase isolated from the venom of Philodryas patagoniensis (Patagonia Green Racer), a South American rear-fanged snake. Herein we demonstrate that patagonfibrase undergoes autolysis at its pH optimum (7.5) and at 37 °C, primarily producing a ∼ 32.6 kDa fragment composed of disintegrin-like and cysteine-rich domains, as identified by mass spectrometry and N-terminal sequencing. The autolysis site for production of this fragment is similar to that observed for metalloproteinases from front-fanged Viperidae snake venoms. In the presence of Ca²⁺, patagonfibrase was only partially autolysed, giving rise mainly to one fragment of ∼ 52.2 kDa. In addition, calcium markedly enhanced the azocaseinolytic activity of patagonfibrase. Our findings contribute to the understanding of the structural and mechanistic bases of this family of metalloenzymes that are widely distributed among snake venoms, demonstrating that important post-translational modifications such as proteolysis can also contribute to the diversity and complexity of proteins found in rear-fanged snake venoms.