title: Role of tyrosine 33 residue for the stabilization of the tetrameric structure of human cytidine deaminase.
abstract: In the present work the effect of a mutation on tyrosine 33 residue (Y33G) of human cytidine deaminase (CDA) was investigated with regard to protein solubility and specific activity. Osmolytes and CDA ligands were used to increase the yield and the specific activity of the protein. The mutant enzyme was purified and subjected to a kinetic characterization and to stability studies. These investigations reinforced the hypothesis that in human CDA the side chain of Y33 is involved in intersubunit interactions with four glutamate residues (E108) forming a double latch that connects each of the two pairs of monomers of the tetrameric CDA.
authors.affiliation: School of Biosciences and Biotechnologies, via Gentile III da Varano, 62032 Camerino (MC), Italy. daniela.micozzi@unicam.it
authors.forename: Daniela
authors.initials: D
authors.lastname: Micozzi
authors.forename: Stefania
authors.initials: S
authors.lastname: Pucciarelli
authors.forename: Francesco M
authors.initials: FM
authors.lastname: Carpi
authors.forename: Stefano
authors.initials: S
authors.lastname: Costanzi
authors.forename: Giampiero
authors.initials: G
authors.lastname: De Sanctis
authors.forename: Valeria
authors.initials: V
authors.lastname: Polzonetti
authors.forename: Paolo
authors.initials: P
authors.lastname: Natalini
authors.forename: Ivano F
authors.initials: IF
authors.lastname: Santarelli
authors.forename: Alberto
authors.initials: A
authors.lastname: Vita
authors.forename: Silvia
authors.initials: S
authors.lastname: Vincenzetti
mesh_terms: D000818:Animals; D015153:Blotting, Western; D020134:Catalytic Domain; D002942:Circular Dichroism; D003564:Cytidine Deaminase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability / Q000187:drug effects; D006801:Humans; D007700:Kinetics; D051379:Mice; D018832:Molecular Chaperones / Q000494:pharmacology; D050505:Mutant Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D058767:Protein Unfolding / Q000187:drug effects; D013329:Structure-Activity Relationship; D013696:Temperature; D014443:Tyrosine / Q000378:metabolism*
chemical_list: D018832:Molecular Chaperones; D050505:Mutant Proteins; D014443:Tyrosine; D003564:Cytidine Deaminase