title: Investigation of the mechanism of interaction between Mannose-binding lectin-associated serine protease-2 and complement C4.
abstract: The interaction between mannose-binding lectin [MBL]-associated serine protease-2 (MASP-2) and its first substrate, C4 is crucial to the lectin pathway of complement, which is vital for innate host immunity, but also involved in a number of inflammatory diseases. Recent data suggests that two areas outside of the active site of MASP-2 (so-called exosites) are crucial for efficient cleavage of C4: one at the junction of the two complement control protein (CCP) domains of the enzyme and the second on the serine protease (SP) domain. Here, we have further investigated the roles of each of these exosites in the binding and cleavage of C4. We have found that both exosites are required for high affinity binding and efficient cleavage of the substrate protein. Within the SP domain exosite, we have shown here that two arginine residues are most important for high affinity binding and efficient cleavage of C4. Finally, we show that the CCP domain exosite appears to play the major role in the initial interaction with C4, whilst the SP domain exosite plays the major role in a secondary conformational change between the two proteins required to form a high affinity complex. This data has provided new insights into the binding and cleavage of C4 by MASP-2, which may be useful in the design of molecules that modulate this important interaction required to activate the lectin pathway of complement.
authors.affiliation: Department of Biochemistry and Molecular Biology, Monash University, Melbourne, Victoria, 3800, Australia.
authors.forename: Nicole
authors.initials: N
authors.lastname: Drentin
authors.affiliation: Department of Biochemistry and Molecular Biology, Monash University, Melbourne, Victoria, 3800, Australia.
authors.forename: Paul
authors.initials: P
authors.lastname: Conroy
authors.affiliation: Department of Biochemistry and Molecular Biology, Monash University, Melbourne, Victoria, 3800, Australia.
authors.forename: Menachem J
authors.initials: MJ
authors.lastname: Gunzburg
authors.affiliation: Department of Biochemistry and Molecular Biology, Monash University, Melbourne, Victoria, 3800, Australia; Department of Biochemistry & Genetics, La Trobe Institute of Molecular Science, La Trobe University, Melbourne, Victoria, 3086, Australia. Electronic address: R.Pike@latrobe.edu.au.
authors.forename: Robert N
authors.initials: RN
authors.lastname: Pike
authors.affiliation: Department of Biochemistry & Genetics, La Trobe Institute of Molecular Science, La Trobe University, Melbourne, Victoria, 3086, Australia.
authors.forename: Lakshmi C
authors.initials: LC
authors.lastname: Wijeyewickrema
mesh_terms: D003181:Complement C4 / Q000378:metabolism*; D055767:Immobilized Proteins / Q000378:metabolism; D007700:Kinetics; D050606:Mannose-Binding Protein-Associated Serine Proteases / Q000737:chemistry / Q000378:metabolism*; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D020349:Surface Plasmon Resonance; D013997:Time Factors
chemical_list: D003181:Complement C4; D055767:Immobilized Proteins; D050505:Mutant Proteins; D050606:Mannose-Binding Protein-Associated Serine Proteases
keywords: C4; Complement; Exosite; Lectin pathway activation; MASP-2; Serine protease