Mass spectrometry analysis of recombinant human ZP3 expressed in glycosylation-deficient CHO cells

Biochemistry
Ming ZhaoJurrien Dean

Abstract

The zona pellucida is an extracellular matrix that mediates taxon-specific fertilization in which human sperm will not bind to mouse eggs. The mouse zona pellucida is composed of three glycoproteins (ZP1, ZP2, ZP3). The primary structure of each has been deduced from the cDNA nucleic acid sequence, and each has been analyzed by mass spectrometry. However, determination of the secondary structure and processing of the human zona proteins have been hampered by the paucity of biological material. To investigate if taxon-specific sperm-egg recognition was ascribable to structural differences in a zona protein required for matrix formation, recombinant human ZP3 was expressed in CHO-Lec3.2.8.1 cells and compared to mouse ZP3. With nearly complete coverage, LC-QTOF mass spectrometry was used to determine the cleavage of an N-terminal signal peptide (amino acids 1-22) and the release of secreted ZP3 from a C-terminal transmembrane domain (amino acids 379-424). The resultant N-terminal glutamine was cyclized to pyroglutamate (pyrGln(23)), and several C-terminal peptides were detected, including one ending at Asn(350). The disulfide bond linkages of eight cysteine residues in the conserved zona domain were ascertained (Cys(46)/Cys(140),...Continue Reading

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Citations

Feb 3, 2012·Cell and Tissue Research·Satish K GuptaNeha Gupta
Apr 8, 2006·BMC Biochemistry·Luca JovinePaul M Wassarman
Sep 14, 2010·Reproductive Biology and Endocrinology : RB&E·Anasua GangulySatish K Gupta
Mar 8, 2011·Journal of Cell Science·Maria Jimenez-Movilla, Jurrien Dean
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Mar 8, 2005·Journal of Mass Spectrometry : JMS
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Jan 29, 2005·The Journal of Biological Chemistry·Tanya HoodbhoyJurrien Dean
Feb 1, 2020·Molecular Reproduction and Development·Eileen FahrenkampLuca Jovine
Nov 17, 2020·The EMBO Journal·Alena StsiapanavaLuca Jovine
Nov 4, 2020·Journal of Assisted Reproduction and Genetics·Dazhi ZhangXianqin Zhang
Apr 18, 2018·Journal of the American Society for Mass Spectrometry·Kelly L WormwoodCostel C Darie

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