Binding of recombinant human proacrosin/acrosin to zona pellucida (ZP) glycoproteins. I. Studies with recombinant human ZPA, ZPB, and ZPC.
OBJECTIVE
To characterize proacrosin/acrosin interaction with isolated zona pellucida (ZP) components.


METHODS
Prospective study.


METHODS
Basic research laboratory.


METHODS
Recombinant proteins derived from human proacrosin (Rec-40, Rec-30, Rec-20, Rec-10, and Rec-6) and from human ZP glycoproteins (rec-hZPA, ZPB, and ZPC).


METHODS
In vitro binding assay developed to assess proacrosin/acrosin-ZP interaction.


METHODS
Zona pellucida glycoprotein binding to proacrosin/acrosin; estimation of binding affinity.


RESULTS
Of all ZP proteins, rec-hZPA demonstrated the highest binding activity toward acrosin (Rec-30) (rec-hZPB: 42% of rec-hZPA; rec-hZPC: 39% of rec-hZPA; P<.0005). Rec-hZPA interaction was disturbed by dextran sulphate (75% inhibition with 10 microM), fucose (67% inhibition with 1.5 microM), and mannose (69% inhibition with 333 mM). Comparing binding activity of proacrosin with other N-terminal acrosin fragments, Rec-40 showed 2.6-3 times higher levels. Moreover, saturable high affinity binding of Rec-40 to ZP components was observed (Kd: 34 nM for rec-hZPA, 38 nM for rec-hZPB, 63 nM for rec-hZPC).


CONCLUSIONS
The rec-hZPA is the major ZP ligand for human proacrosin/acrosin. The interaction involves mannosyl, fucosyl, and sulfated glycans. Binding sites for rec-hZP would be located both at the N- and C-terminus of proacrosin, revealing a key role of the proenzyme in the interaction.
Furlong
Laura I
LI
Instituto de Biología y Medicina Experimental-CONICET-UBA, Buenos Aires, Argentina. mhvaz@dna.uba.ar
Harris
Jeffrey D
JD
Vazquez-Levin
Mónica H
MH
D000176:Acrosin / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D000818:Animals; D001665:Binding Sites / Q000502:physiology; D016466:CHO Cells; D006224:Cricetinae; D004527:Egg Proteins / Q000378:metabolism*; D004792:Enzyme Precursors / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D006801:Humans; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism*; D011446:Prospective Studies; D011485:Protein Binding / Q000502:physiology; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D015044:Zona Pellucida / Q000201:enzymology* / Q000378:metabolism / Q000502:physiology; D000072216:Zona Pellucida Glycoproteins
D004527:Egg Proteins; D004792:Enzyme Precursors; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; C082833:ZP4 protein, human; D000072216:Zona Pellucida Glycoproteins; C022438:proacrosin; D000176:Acrosin