title: Isolation and characterization of Bradykinin potentiating peptides from Agkistrodon bilineatus venom.
abstract: BACKGROUND
  Snake venom is a source of many pharmacologically important molecules. Agkistrodon bilineatus commonly known as Cantil, is spread over Central America particularly in Mexico and Costa Rica. From the venom of Agkistrodon bilineatus we have isolated and characterised six hypotensive peptides, and two bradykinin inhibitor peptides. The IC-50 value of four synthesized peptides was studied, towards angiotensin converting enzyme, in order to study the structure-function relationship of these peptides.
  
  
  RESULTS
  The purification of the peptides was carried out by size exclusion chromatography, followed by reverse phase chromatography. Sequences of all peptides were determined applying MALDI-TOF/TOF mass spectrometry. These hypotensive peptides bear homology to bradykinin potentiating peptides and venom vasodilator peptide. The peptide with m/z 1355.53 (M + H)(+1), and the corresponding sequence ZQWAQGRAPHPP, we identified for the first time. A precursor protein containing a fragment of this peptide was reported at genome level, (Uniprot ID P68515), in Bothrops insularis venom gland. These proline rich hypotensive peptides or bradykinin potentiating peptides are usually present in the venom of Crotalinae, and exhibit specificity in binding to the C domain of somatic angiotensin converting enzyme. Four of these hypotensive peptides, were selected and synthesized to obtain the required quantity to study their IC50 values in complex with the angiotensin converting enzyme. The peptide with the sequence ZLWPRPQIPP displayed the lowest IC50 value of 0.64 μM. The IC50 value of the peptide ZQWAQGRAPHPP was 3.63 μM.
  
  
  CONCLUSIONS
  The canonical snake venom BPPs classically display the IPP motif at the C-terminus. Our data suggest that the replacement of the highly conserved hydrophobic isoleucine by histidine does not affect the inhibitory activity, indicating that isoleucine is not mandatory to inhibit the angiotensin converting enzyme. The evaluation of IC 50 values show that the peptide with basic pI value exhibits a lower IC 50 value.
authors.affiliation: Department of Chemistry, University of Engineering & Technology, G.T. Road, 54890 Lahore, Pakistan.
authors.forename: Aisha
authors.initials: A
authors.lastname: Munawar
authors.affiliation: Department of Chemistry, University of Engineering & Technology, G.T. Road, 54890 Lahore, Pakistan.
authors.forename: Anum
authors.initials: A
authors.lastname: Zahid
authors.affiliation: Biochemistry Division, Department of Chemistry, Faculty of Science, Mansoura University, Mansoura, Egypt.
authors.forename: Amr
authors.initials: A
authors.lastname: Negm
authors.affiliation: Botany Division, Institute of Pure and Applied Biology, Bahauddin Zakariya University, 60800 Multan, Pakistan.
authors.forename: Ahmed
authors.initials: A
authors.lastname: Akrem
authors.affiliation: Biotechnology Centre, Instituto de Pesquisas Energeticas e Nucleares, Avenida Lineu Prestes, 2242 São Paulo, Brazil.
authors.forename: Patrick
authors.initials: P
authors.lastname: Spencer
authors.affiliation: Department of Chemistry, University of Hamburg, 20146, Martin-Luther-King Platz 6, 22607 Hamburg, Germany ; Institute of Biochemistry and Molecular Biology, Martin-Luther-King Platz 6, 20146 Hamburg, Germany ; Laboratory for Structural Biology of Infection and Inflammation, DESY, Build. 22a Notkestr. 85, 22603 Hamburg, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Betzel
keywords: Angiotensin converting enzyme; Hypotension; MALDI/TOF-TOF; Peptides; Size exclusion chromatography; Snake venom