Structural Characterization of Recombinant Soluble Rat Neuroligin 1: Mapping of Secondary Structure and Glycosylation by Mass Spectrometry
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- 作者：Ross C. Hoffman ; Lori L. Jennings ; Igor Tsigelny ; Davide Comoletti ; Robyn E. Flynn ; Thomas C. Sudhof ; and Palmer Taylor
- 刊名：Biochemistry
- 出版年：2004
- 出版时间：February 17, 2004
- 年：2004
- 卷：43
- 期：6
- 页码：1496 - 1506
- 全文大小：351K
- 年卷期：v.43,no.6(February 17, 2004)
- ISSN：1520-4995
文摘
Neuroligins (NLs) are a family of transmembrane proteins that function in synapse formationand/or remodeling by interacting with 
fchars/beta2.gif" BORDER=0 ALIGN="middle">-neurexins (fchars/beta2.gif" BORDER=0 ALIGN="middle">-NXs) to form heterophilic cell adhesions. The largeN-terminal extracellular domain of NLs, required for fchars/beta2.gif" BORDER=0 ALIGN="middle">-NX interactions, has sequence homology to thefchars/alpha.gif" BORDER=0>/fchars/beta2.gif" BORDER=0 ALIGN="middle"> hydrolase fold superfamily of proteins. By peptide mapping and mass spectrometric analysis of asoluble recombinant form of NL1, several structural features of the extracellular domain have beenestablished. Of the nine cysteine residues in NL1, eight are shown to form intramolecular disulfide bonds.Disulfide pairings of Cys 117 to Cys 153 and Cys 342 to Cys 353 are consistent with disulfide linkagesthat are conserved among the family of fchars/alpha.gif" BORDER=0>/fchars/beta2.gif" BORDER=0 ALIGN="middle"> hydrolase proteins. The disulfide bond between Cys 172 andCys 181 occurs within a region of the protein encoded by an alternatively spliced exon. The disulfidepairing of Cys 512 and Cys 546 in NL1 yields a structural motif unique to the NLs, since these residuesare highly conserved. The potential N-glycosylation sequons in NL1 at Asn 109, Asn 303, Asn 343, andAsn 547 are shown occupied by carbohydrate. An additional consensus sequence for N-glycosylation atAsn 662 is likely occupied. Analysis of N-linked oligosaccharide content by mass matching paradigmsreveals significant microheterogeneous populations of complex glycosyl moieties. In addition, O-linkedglycosylation is observed in the predicted stalk region of NL1, prior to the transmembrane spanning domain.From predictions based on sequence homology of NL1 to acetylcholinesterase and the molecular featuresof NL1 established from mass spectrometric analysis, a novel topology model for NL three-dimensionalstructure has been constructed.
fchars/beta2.gif" BORDER=0 ALIGN="middle">-neurexins (fchars/beta2.gif" BORDER=0 ALIGN="middle">-NXs) to form heterophilic cell adhesions. The largeN-terminal extracellular domain of NLs, required for fchars/beta2.gif" BORDER=0 ALIGN="middle">-NX interactions, has sequence homology to thefchars/alpha.gif" BORDER=0>/fchars/beta2.gif" BORDER=0 ALIGN="middle"> hydrolase fold superfamily of proteins. By peptide mapping and mass spectrometric analysis of asoluble recombinant form of NL1, several structural features of the extracellular domain have beenestablished. Of the nine cysteine residues in NL1, eight are shown to form intramolecular disulfide bonds.Disulfide pairings of Cys 117 to Cys 153 and Cys 342 to Cys 353 are consistent with disulfide linkagesthat are conserved among the family of fchars/alpha.gif" BORDER=0>/fchars/beta2.gif" BORDER=0 ALIGN="middle"> hydrolase proteins. The disulfide bond between Cys 172 andCys 181 occurs within a region of the protein encoded by an alternatively spliced exon. The disulfidepairing of Cys 512 and Cys 546 in NL1 yields a structural motif unique to the NLs, since these residuesare highly conserved. The potential N-glycosylation sequons in NL1 at Asn 109, Asn 303, Asn 343, andAsn 547 are shown occupied by carbohydrate. An additional consensus sequence for N-glycosylation atAsn 662 is likely occupied. Analysis of N-linked oligosaccharide content by mass matching paradigmsreveals significant microheterogeneous populations of complex glycosyl moieties. In addition, O-linkedglycosylation is observed in the predicted stalk region of NL1, prior to the transmembrane spanning domain.From predictions based on sequence homology of NL1 to acetylcholinesterase and the molecular featuresof NL1 established from mass spectrometric analysis, a novel topology model for NL three-dimensionalstructure has been constructed.