Login (DCU Staff Only)
Login (DCU Staff Only)

DORAS | DCU Research Repository

Explore open access research and scholarly works from DCU

Advanced Search

The purification and characterisation of novel dipeptidyl peptidase IV-like activity from bovine serum

Buckley, Seamus J., Collins, Patrick and O'Connor, Brendan orcid logoORCID: 0000-0002-6857-1614 (2004) The purification and characterisation of novel dipeptidyl peptidase IV-like activity from bovine serum. International Journal of Biochemistry & Cell Biology, 36 (7). pp. 1281-1296. ISSN 1357-2725

Abstract
The discovery of a potentially novel proline-specific peptidase from bovine serum is presented which is capable of cleaving the dipeptidyl peptidase IV (DPIV) substrate Gly-Pro-MCA. The enzyme was isolated and purified with the use of Phenyl Sepharose Hydrophobic Interaction, Sephacryl S300 Gel Filtration, and Q-Sephacryl Anion Exchange, producing an overall purification factor of 257. SDS PAGE resulted in a monomeric molecular mass of 158 kDa while Size Exclusion Chromatography generated a native molecular mass of 328 kDa. The enzyme remained active over a broad pH range with a distinct preference for a neutral pH range of 7-8.5. Chromatofocusing and Isoelectric Focusing revealed the enzyme’s isoelectric point to be 4.74. DPIV-like activity was not inhibited by serine protease inhibitors but was by the metallo-protease inhibitors, the phenanthrolines. The enzyme was also partially inhibited by Bestatin. Substrate Specificity studies proved that the enzyme is capable of sequential cleavage of bovine β- Casomorphin and Substance P. The peptidase cleaved the standard DPIV substrate, Gly-Pro-MCA with a KM of 38.4 μM, while Lys-Pro-MCA was hydrolysed with a KM of 103 μM. The DPIV- like activity was specifically inhibited by both Diprotin A and B, non-competitively, generating a Ki of 1.4x10-4 M for both inhibitors. Ile-Thiazolidide and Ile-Pyrrolidide both inhibited competitively with an inhibition constant of 3.7x10-7 M and 7.5x10-7 M respectively. It is concluded that bovine serum DPIV-like activity share many biochemical properties with DPIV and DPIV-like enzymes but not exclusively, suggesting that the purified peptidase may play an important novel role in bioactive oligopeptide degradation.
Metadata
Item Type:Article (Published)
Refereed:Yes
Subjects:Biological Sciences > Biotechnology
Biological Sciences > Biochemistry
Biological Sciences > Enzymology
DCU Faculties and Centres:DCU Faculties and Schools > Faculty of Science and Health > School of Biotechnology
Research Initiatives and Centres > Irish Separation Science Cluster (ISSC)
Publisher:Elsevier
Official URL:http://dx.doi.org/10.1016/j.biocel.2003.02.001
Copyright Information:© 2004 Elsevier
Use License:This item is licensed under a Creative Commons Attribution-NonCommercial-Share Alike 3.0 License. View License
Funders:Enterprise Ireland, Health Research Board
ID Code:17813
Deposited On:28 Feb 2013 14:41 by Brendan O'connor . Last Modified 18 Oct 2018 13:25
Documents

Full text available as:

[thumbnail of DPIVpaper_'04.pdf]
Preview
PDF - Requires a PDF viewer such as GSview, Xpdf or Adobe Acrobat Reader
200kB
Downloads

Downloads

Downloads per month over past year

Archive Staff Only: edit this record