A novel and highly efficient purification procedure for native human dipeptidyl peptidase 3 from human blood cell lysate

Abstract

Dipeptidyl amino-peptidase 3 (DPP3) is an aminopeptidase involved in peptide degradation, including hormone peptides as angiotensin II and enkephalins. DPP3 plasma activity increases in septic patients and correlates with mortality risk. However, the exact physiological role of DPP3 remains unclear and animal studies are necessary to reveal the function of DPP3 in vivo . To this demand, we developed a two-step purification procedure for isolation of native human DPP3 from blood cell lysate (BCL) that is suitable for in vivo applications. With the use of monoclonal antibodies coupled to beads in combination with an ion-exchange chromatography, we recovered 68% of human DPP3 activity from BCL with a purity of ≥ 95%. Purified human DPP3 was assayed for activity and protein concentration using recently published DPP3-activity- and immunoassays. Additionally, protein stability and storage in relevant buffers were tested. Our results provide a promising strategy for fast and efficient isolation of human DPP3. The purified human DPP3 represents the native state of DPP3, suitable for future in vivo applications to investigate the physiological role of DPP3 and its involvement in pathophysiological conditions.

Circulating dipeptidyl peptidase 3 is a myocardial depressant factor: dipeptidyl peptidase 3 inhibition rapidly and sustainably improves haemodynamics

“…In order to assess the direct effects of cDPP3 on cardiovascular function and kidney haemodynamics, purified native human DPP3 (hDPP3) was intravenously injected into healthy anaesthetized mice ( Figure A ). Injected hDPP3 had a half‐life of about 20 min ( Figure B ), and hDPP3 concentrations returned to low values at 120 min, as previously observed in healthy rats .…”

“…Native human DPP3 (hDPP3) was purified from human blood cells as previously described and injected into 3‐month‐old male C56Bl/6 mice (0.55 mg/kg). Of note, identity between human DPP3 (accession number NP_569710) and mouse DPP3 (accession number NP_001347640) is 93% and similarity is 96% over the entire length of the proteins (737 amino acids).…”

Quality Control of Jinhua Ham from the Influence between Proteases Activities and Processing Parameters: A Review

“…DPP III reaches its maximum activity at a pH of 8.0 and a temperature of . Its activity decreases rapidly under acidic conditions and decreases significantly when the optimum temperature is exceeded [ 38 ]. DPP IV is a dipeptidase that is second only to DPP I in activity and it has a positive effect on the proteolysis.…”

The emerging role of dipeptidyl peptidase 3 in pathophysiology

“…To further validate the findings of the shock cohort analysis, efficient methods for native DPP3 purification [106] and animal models to study the pathophysiological role of DPP3 were developed. It was shown that (a) DPP3 intravenous injection leads to reduced cardiac and renal function; (b) high DPP3 blood levels induced by acute cardiac stress or septic cardiomyopathy can be inhibited by a humanized monoclonal anti‐cDPP3 IgG1 antibody (anti‐cDPP3‐mAb), which normalized cardiac function, restored hemodynamic stability and reduced mortality [47].…”