Expression and purification of the recombinant His-tagged GST-CD38 fusion protein using the baculovirus/insect cell expression system.
CD38 is a type II transmembrane glycoprotein found in myriad mammalian tissues and cell types. It is known for its involvement in the metabolism of cyclic ADP-ribose and nicotinic acid adenine dinucleotide phosphate, two nucleotides with calcium mobilizing activity independent of inositol trisphosphate. CD38 itself has been shown to have clinical significance in certain diseases with possible utilization in diagnostic and prognostic applications. Previous studies on several autoimmune diseases have shown the usefulness of recombinant CD38 protein expressed from Escherichia coli and Pichia pastoris in the detection of autoantibodies to CD38 via Western blot and ELISA. In this study, we produced a 6 x His-tagged GST-CD38 fusion protein using a recombinant baculovirus/insect cell expression technique that was purified as a soluble protein. The fusion protein was purified to homogeneity by affinity and gel filtration chromatography steps. It has an apparent molecular mass of 56 kDa on SDS-PAGE gel stained with Coomassie blue and was recognized on Western blots by antibodies against human CD38 as well as the polyhistidine tag. Peptide mass fingerprinting analysis confirmed the identity of human CD38 in the fusion protein.
Khoo
Keng Meng
KM
Department of Rheumatology, Allergy, and Immunology, Tan Tock Seng Hospital, 11 Jalan Tan Tock Seng, Singapore 308433, Singapore. keng_meng_khoo@ttsh.com.sg
Chang
Chan Fong
CF
Schubert
Jeffrey
J
Wondrak
Ewald
E
Chng
Hiok Hee
HH
D036541:ADP-ribosyl Cyclase / Q000235:genetics* / Q000276:immunology; D051997:ADP-ribosyl Cyclase 1; D000818:Animals; D000915:Antibody Affinity; D015703:Antigens, CD / Q000235:genetics* / Q000276:immunology; D016367:Baculoviridae; D002460:Cell Line; D003001:Cloning, Molecular / Q000379:methods*; D018076:DNA, Complementary / Q000235:genetics; D005982:Glutathione Transferase / Q000235:genetics*; D006639:Histidine; D006801:Humans; D007313:Insecta / Q000166:cytology; D008562:Membrane Glycoproteins; D015335:Molecular Probes; D010449:Peptide Mapping; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification*
D015703:Antigens, CD; D018076:DNA, Complementary; D008562:Membrane Glycoproteins; D015335:Molecular Probes; D011993:Recombinant Fusion Proteins; D006639:Histidine; D005982:Glutathione Transferase; D036541:ADP-ribosyl Cyclase; C498154:CD38 protein, human; D051997:ADP-ribosyl Cyclase 1