Use of isothermal titration calorimetry to investigate and identify novel peptide substrates for prolyl carboxypeptidase: a technology comparison

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00:00:00 Use of isothermal titration calorimetry to investigate and identify novel peptide substrates for prolyl carboxypeptidase: a technology comparison
00:01:23 Use of isothermal titration calorimetry to investigate and identify novel peptide substrates for propyl carboxypeptidase.
00:01:40 Introduction
00:02:00 Introduction
00:02:13 Introduction
00:02:17 Introduction
00:02:27 Introduction
00:02:39 Introduction
00:02:57 Introduction
00:03:08 Aims
00:03:14 Aims
00:03:17 Aims
00:03:22 Aims
00:03:26 Aims
00:03:42 Aims
00:04:05 Aims
00:04:17 Isothermal Titration Calorimetry (ITC)
00:04:38 Isothermal Titration Calorimetry (ITC)
00:04:45 Isothermal Titration Calorimetry (ITC)
00:04:54 Isothermal Titration Calorimetry (ITC)
00:05:02 Isothermal Titration Calorimetry (ITC)
00:05:31 Isothermal Titration Calorimetry (ITC)
00:05:37 Isothermal Titration Calorimetry (ITC)
00:05:41 Isothermal Titration Calorimetry (ITC)
00:05:44 Isothermal Titration Calorimetry (ITC)
00:05:53 Isothermal Titration Calorimetry (ITC)
00:06:15 Isothermal Titration Calorimetry (ITC)
00:06:20 Isothermal Titration Calorimetry (ITC)
00:06:23 Isothermal Titration Calorimetry (ITC)
00:06:26 Isothermal Titration Calorimetry (ITC)
00:06:32 Isothermal Titration Calorimetry (ITC)
00:06:36 Isothermal Titration Calorimetry (ITC)
00:06:42 Isothermal Titration Calorimetry (ITC)
00:06:50 Isothermal Titration Calorimetry (ITC)
00:07:05 Isothermal Titration Calorimetry (ITC)
00:07:31 Isothermal Titration Calorimetry (ITC)
00:07:38 Isothermal Titration Calorimetry (ITC)
00:07:51 Isothermal Titration Calorimetry (ITC)
00:07:55 Isothermal Titration Calorimetry (ITC)
00:08:00 Isothermal Titration Calorimetry (ITC)
00:08:15 1. Determine peptide and enzyme concentrations
00:08:32 1. Determine peptide and enzyme concentrations
00:08:38 1. Determine peptide and enzyme concentrations
00:08:57 1. Determine peptide and enzyme concentrations
00:09:05 1. Determine peptide and enzyme concentrations
00:09:11 1. Determine peptide and enzyme concentrations
00:09:18 1. Determine peptide and enzyme concentrations
00:09:34 1. Determine peptide and enzyme concentrations
00:09:38 1. Determine peptide and enzyme concentrations
00:09:41 1. Determine peptide and enzyme concentrations
00:09:46 1. Determine peptide and enzyme concentrations
00:09:51 1. Determine peptide and enzyme concentrations
00:09:58 1. Determine peptide and enzyme concentrations
00:10:02 1. Determine peptide and enzyme concentrations
00:10:16 1. Determine peptide and enzyme concentrations
00:10:27 2. Determine total molar enthalpy
00:10:43 2. Determine total molar enthalpy
00:10:53 2. Determine total molar enthalpy
00:11:02 2. Determine total molar enthalpy
00:11:07 2. Determine total molar enthalpy
00:11:30 2. Determine total molar enthalpy
00:11:36 2. Determine total molar enthalpy
00:11:40 2. Determine total molar enthalpy
00:11:51 2. Determine total molar enthalpy
00:11:59 2. Determine total molar enthalpy
00:12:05 2. Determine total molar enthalpy
00:12:29 3. Perform ITC experiment
00:12:39 3. Perform ITC experiment
00:12:44 3. Perform ITC experiment
00:12:52 3. Perform ITC experiment
00:13:11 Results
00:13:15 1. Hydrolysis of angiotensin III by rPRCP
00:13:22 1. Hydrolysis of angiotensin III by rPRCP
00:13:41 1. Hydrolysis of angiotensin III by rPRCP
00:13:51 1. Hydrolysis of angiotensin III by rPRCP
00:14:02 1. Hydrolysis of angiotensin III by rPRCP
00:14:16 2. Hydrolysis of (pyr)-apelin-13 by rPRCP
00:14:23 2. Hydrolysis of (pyr)-apelin-13 by rPRCP
00:14:30 2. Hydrolysis of (pyr)-apelin-13 by rPRCP
00:14:37 2. Hydrolysis of (pyr)-apelin-13 by rPRCP
00:14:48 2. Hydrolysis of (pyr)-apelin-13 by rPRCP
00:14:57 2. Hydrolysis of (pyr)-apelin-13 by rPRCP
00:15:09 3. Method comparison
00:16:02 3. Method comparison
00:16:30 3. Method comparison
00:17:09 Conclusion
00:17:17 Conclusion
00:17:23 Conclusion
00:17:30 Conclusion
00:17:49 Acknowledgements
00:18:09 Acknowledgements
00:18:17 Thank you for your attentionAny questions?
00:20:11 Untitled
00:21:21 Thank you for your attentionAny questions?
00:21:46 Contact Information
The search for novel peptide substrates for proline-specific peptidases has been hampered by a lack of suitable assays. Existing technologies are often too expensive, labor-intensive or require derivatization and separation of the reaction products. The purpose of this study was to validate the use of isothermal titration calorimetry (ITC) to study enzyme kinetics, in comparison to more conventional methods (e.g. MALDI-TOF mass spectrometry, quantitation of released C-terminal Phe using RP-HPLC). The study led to the discovery of a novel substrate for prolyl carboxypeptidase, an enzyme involved in body weight regulation.