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Serine protease identification ( in vitro ) and molecular structure predictions ( in silico ) from a phytopathogenic fungus, Alternaria solani
Author(s) -
Chandrasekaran Murugesan,
Chandrasekar Raman,
Sa Tongmin,
Sathiyabama Muthukrishnan
Publication year - 2014
Publication title -
journal of basic microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.58
H-Index - 54
eISSN - 1521-4028
pISSN - 0233-111X
DOI - 10.1002/jobm.201300433
Subject(s) - serine , proteases , serine protease , biochemistry , protease , stereochemistry , isoelectric point , ramachandran plot , biology , alternaria solani , chemistry , protein structure , enzyme , botany , fungicide
Serine proteases are involved in an enormous number of biological processes. The present study aims at characterizing three‐dimensional (3D) molecular architecture of serine proteases from early blight pathogen, Alternaria solani that are hypothesized to be markers of phytopathogenicity. A serine protease was purified to homogeneity and MALDI‐TOF‐MS/MS analysis revealed that protease produced by A. solani belongs to alkaline serine proteases (AsP). AsP is made up of 403 amino acid residues with molecular weight of 42.1 kDa (Isoelectric point – 6.51) and its molecular formula was C 1859 H 2930 N 516 O 595 S 4 . AsP structure model was built based on its comparative homology with serine protease using the program, MODELER. AsP had 16 β‐sheets and 10 α‐helices, with Ser 350 (G347–G357), Asp 158 (D158–H169), and His 193 (H193–G203) in separate turn/coil structures. Biological metal binding region situated near 6th‐helix and His 193 residue is responsible for metal binding site. Also, calcium ion (Ca 2+ ) is coordinated by the carboxyl groups of Lys 84 , Ile 85 , Lys 86 , Asp 87 , Phe 88 , Ala 89 , Ala 90 (K84–A90) for first Ca 2+ binding site and carbonyl oxygen atom of Lys 244 , Gly 245 , Arg 246 , Thr 247 , Lys 248 , Lys 249 , and Ala 250 (K244–A250), for second Ca 2+ binding site. Moreover, Ramachandran plot analysis of protein residues falling into most favored secondary structures were determined (83.3%). The predicted molecular 3D structural model was further verified using PROCHECK, ERRAT, and VADAR servers to confirm the geometry and stereo‐chemical parameters of the molecular structural design. The functional analysis of AsP 3D molecular structure predictions familiar in the current study may provide a new perspective in the understanding and identification of antifungal protease inhibitor designing.