Circulin-B

Circulin A and B are members of a family of macrocyclic peptides, originally isolated from the tropical tree Chassalia parvifolia, that have been shown to display anti-HIV activity. Complete structural elucidation of these highly constrained peptides was difficult due to their cyclic amide backbone and the presence of six disulfide-linked cysteines. In the present study, the disulfide pairing motif of circulin A and circulin B was determined. Since the circulins were resistant to enzymatic proteolysis, cysteine residue pairings were identified by analysis of the complex mixture of cleavage products that resulted from partial acid hydrolysis of the native peptides. Combined utilization of HPLC, fast atom bombardment mass spectrometry and peptide recognition software ("F-MASS" and "F-LINK" programs) were employed to identify the cleavage products. Thus, we were able to unambiguously identify the disulfide linkage pattern in circulin A and circulin B as Cys1-Cys4, Cys2-Cys5 and Cys3-Cys6, where the numbers on the cystine residues refer to their respective order in the peptides.

Computational methods have refined the mode of peptide drug designing to a new plateau recently. Circulin, a 30 residue natural plant polypeptide acts as a plant defensive peptide. Additional to its antimicrobial activity it also possesses an inhibitory cytopathic effect on the replication of human immunodeficiency virus (HIV). Stable Circulin can be a functionally able template for scaffolding peptide based drugs. Hence, structural stability of Chassalia parvifolia, Circulin A (1BH4), and Circulin B (2ERI) was computationally investigated. From this analysis, the stability favored toward Circulin B which was supported by various parameters such as intra-molecular interactions (61), secondary structure, hydrophobicity (67.34%), root mean square deviation (2.64Å), root mean square fluctuation (0.08Å), radius of gyration (8.96Å), ovality (3.49), angular deviation (73.6%), surface area (both polar and non-polar), hydrogen bond distribution (11.94), and disulphide bond distances. Further, the functional activity calculated in terms of membrane associated free energy (-4.10 kcal/mol) also favored Circulin B. Hence, Circulin B could be proposed as the best template for scaffolding antimicrobial as well as antiviral (HIV) peptide based drug design. The obtained computational data can aid experimental biologists to successfully produce stable therapeutic peptides from natural resources reducing erroneous wastage of monetary sources and time.