Isoenzymatic forms of human cytidine deaminase.
Cytidine deaminase (CDA) purified from human placenta revealed the presence of five isoenzymatic forms that differ only in their isoelectric point. Since human cytidine deaminase exists in two variants (CDA 1 and CDA 2) with a non-conservative amino acid substitution at codon 27, in this work we demonstrate that these two variants may combine together in vitro, giving five CDA isoforms as observed in vivo from human placenta. For this purpose, each of the two forms of CDA was purified close to homogeneity and dissociated into monomers in the presence of a small amount of sodium dodecyl sulfate as a dissociating agent. The monomers were mixed together and subjected to anion-exchange chromatography and to chromatofocusing analysis in order to visualize the formation of the five isoforms. Furthermore, for both CDA 1 and CDA 2 some substrates and inhibitors of CDA were assayed, with the aim of demonstrating different kinetic behavior between the two natural variants.
Vincenzetti
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Dipartimento di Scienze Veterinarie, Dipartimento di Scienze Morfologiche e Biochimiche Comparate and Dipartimento di Scienze Chimiche, Università di Camerino, Via Circonvallazione 93/95, 62024 Matelica (MC), Italy.
Mariani
P L
PL
Cammertoni
N
N
Polzonetti
V
V
Natalini
P
P
Quadrini
B
B
Volpini
R
R
Vita
A
A
D000837:Anion Exchange Resins / Q000494:pharmacology; D002845:Chromatography; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D003062:Codon; D003564:Cytidine Deaminase / Q000737:chemistry* / Q000302:isolation & purification*; D018076:DNA, Complementary / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D007525:Isoelectric Focusing; D007700:Kinetics; D010920:Placenta / Q000201:enzymology; D015202:Protein Engineering / Q000379:methods; D020033:Protein Isoforms; D011994:Recombinant Proteins / Q000737:chemistry; D012117:Resins, Synthetic; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry
D000837:Anion Exchange Resins; D003062:Codon; D018076:DNA, Complementary; D020033:Protein Isoforms; D011994:Recombinant Proteins; D012117:Resins, Synthetic; D012967:Sodium Dodecyl Sulfate; C042855:Mono Q; D003564:Cytidine Deaminase