Applied Biochemistry and Biotechnology

, Volume 171, Issue 2, pp 366–381 | Cite as

Buffalo Colostrum β-lactoglobulin Inhibits VEGF-Induced Angiogenesis by Interacting with G protein-Coupled Receptor Kinase

  • Rohit A. Chougule
  • Shilpa P.
  • Bharathi P. Salimath
  • Aparna H. Sosalegowda
Article
First Online:
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Accepted:

Abstract

β-lactoglobulin (β-lg), a major whey protein was purified and characterised from buffalo colostrum. The in silico analysis of the tryptic peptides based on LC-CID-MS/MS facilitated the identification of protein as β-lg. The sequences IIVTQ f[1–5] and LSFNPTQLEEQCHV f(149–162) of m/z 933+ and 8512+ were found to match N- and C-extreme of β-lg while IDALNENK f(84–91) and TPEVDDEALEKFDK f(125–138) sequences deduced for m/z 916+ and 8182+ were in compliance to buffalo milk β-lg. Considering the sequence similarity of β-lg to glycodelin, a proven angiogenic protein, similar role for β-lg from buffalo colostrum (BLG-col) was examined. Interestingly, BLG-col exhibited anti-angiogenic activity by potently inhibiting cell proliferation, micro-vessel sprouting, cell migration and tube formation of human umbilical vein endothelial cells (HUVECs) in a dose-dependent manner but having varied effect on Ehrlich ascites tumor cells, MCF-7, MDA-MB 435 and MDA-MB 231 cell lines. The anti-angiogenic potential of BLG-col was found to be vascular endothelial growth factor mediated. The immunolocalisation of BLG-col on the cell surface of HUVECs evidenced using FITC-labelled β-lg antibody indicated its extra-cellular binding. Furthermore, BLG-col interacting HUVEC membrane protein (64 kDa) was detected by immunoblot and its identity was established by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry analysis, which showed peptide sequence homology to G protein-coupled receptor kinase 4.

Keywords

Buffalo colostrum β-lactoglobulin Anti-angiogenic activity GRK 4 
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Notes

Acknowledgements

We thank University Grant Commission, New Delhi, India for sanction of grant (F31-294/2005-06) to undertake this investigation and Rohit A. Chougule thank ICMR for award of SRF.

Supplementary material

12010_2013_344_MOESM1_ESM.doc (1.4 mb)
ESM 1 (DOC 1471 kb)

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Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Rohit A. Chougule
    • 1
  • Shilpa P.
    • 1
  • Bharathi P. Salimath
    • 1
  • Aparna H. Sosalegowda
    • 1
  1. 1.Department of Studies in Biotechnology, ManasagangotriUniversity of MysoreMysoreIndia

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