Identifying the structure of the active sites of human recombinant prolidase.
In this paper we provide a detailed biochemical and structural characterization of the active site of recombinant human prolidase, a dimeric metalloenzyme, whose misfunctioning causes a recessive connective tissue disorder (prolidase deficiency) characterized by severe skin lesions, mental retardation and respiratory tract infections. It is known that the protein can host two metal ions in the active site of each constituent monomer. We prove that two different kinds of metals (Mn and Zn) can be simultaneously present in the protein active sites with the protein partially maintaining its enzymatic activity. Structural information extracted from X-ray absorption spectroscopy measurements have been used to yield a full reconstruction of the atomic environment around each one of the two monomeric active sites. In particular, as for the metal ion occupation configuration of the recombinant human prolidase, we have found that one of the two active sites is occupied by two Zn ions and the second one by one Zn and one Mn ion. In both dinuclear units a histidine residue is bound to a Zn ion.
Dipartimento di Biochimica, Sezione Medicina e Farmacia, Università di Pavia, Pavia, Italy.
Roberta
R
Besio
Stefania
S
Alleva
Antonella
A
Forlino
Anna
A
Lupi
Carlo
C
Meneghini
Velia
V
Minicozzi
Antonella
A
Profumo
Francesco
F
Stellato
Ruggero
R
Tenni
Silvia
S
Morante
D001665:Binding Sites / Q000187:drug effects*; D020134:Catalytic Domain; D004150:Dipeptidases / Q000737:chemistry*; D006801:Humans; D007477:Ions; D008345:Manganese / Q000737:chemistry; D008667:Metalloproteins / Q000737:chemistry*; D008670:Metals / Q000737:chemistry; D056732:Prolidase Deficiency / Q000378:metabolism*; D011487:Protein Conformation; D013379:Substrate Specificity; D015032:Zinc / Q000737:chemistry
D007477:Ions; D008667:Metalloproteins; D008670:Metals; D008345:Manganese; D004150:Dipeptidases; C022449:proline dipeptidase; D015032:Zinc