Purification of NAD+ glycohydrolase enzyme from erythrocyte membrane

Objective: In this study our object was the purification of the erythrocyte membrane bound NAD glycohydrolase of the CD38 surface antigen that is involved in the lymphocyte differentiation and activation and is also found on the erythrocyte membrane. We think that the purification of the enzyme will elucidate the structure and the activity of the CD38 and consequently the maturation, aging and death of the hemopoietic cells and the pathologic mechanisms involved in this process. Material and Methods: Fresh blood samples from the healthy individuals were obtained by anticoagulated with heparin. Solubilized eythrocyte ghosts were purified utilizing hydroxylapatite resin, Sephadex G-100 (gel filtration chromatography), Cu++ -bound IDA (iminodiacetic acid), chromatography, octyl agarose, DEAE cellulose column chromatography steps, respectively. Results: A protein band with a molecular weight 45 000 Dalton was obtained in the SDS-PAGE analysis after the purification. Conclusion: Membrane bound NAD+ glycohydrolase activity after purification was found 257 times higher than the initial erythrocyte ghost yield. The yield of the purification was 1.4%. Copyright © 2010 by Türkiye Klinikleri.