Effects of the N-Linked Glycans on the 3D Structure of the Free -Subunit of Human Chorionic Gonadotropin
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- 作者：Paul J. A. Erbel ; Yasmin Karimi-Nejad ; J. Albert van Kuik ; Rolf Boelens ; Johannis P. Kamerling ; and Johannes F. G. Vliegenthart
- 刊名：Biochemistry
- 出版年：2000
- 出版时间：May 23, 2000
- 年：2000
- 卷：39
- 期：20
- 页码：6012 - 6021
- 全文大小：775K
- 年卷期：v.39,no.20(May 23, 2000)
- ISSN：1520-4995
文摘
To gain insight into intramolecular carbohydrate-protein interactions at the molecular level,the solution structure of differently deglycosylated variants of the 
-subunit of human chorionicgonadotropin have been studied by NMR spectroscopy. Significant differences in chemical shifts andNOE intensities were observed for amino acid residues close to the carbohydrate chain at Asn78 upondeglycosylation beyond Asn78-bound GlcNAc. As no straightforward strategy is available for the calculationof the NMR structure of intact glycoproteins, a suitable computational protocol had to be developed. Tothis end, the X-PLOR carbohydrate force field designed for structure refinement was extended and modified.Furthermore, a computational strategy was devised to facilitate successful protein folding in the presenceof extended glycans during the simulation. The values for 
and 
dihedral angles of the glycosidiclinkages of the oligosaccharide core fragments GlcNAc2(
1-4)GlcNAc1 and Man3(1-4)GlcNAc2 arerestricted to a limited range of the broad conformational energy minima accessible for free glycans. Thisdemonstrates that the protein core affects the dynamic behavior of the glycan at Asn78 by steric hindrance.Reciprocally, the NMR structures indicate that the glycan at Asn78 affects the stability of the proteincore. The backbone angular order parameters and displacement data of the generated conformers displayespecially for the -turn 20-23 a decreased structural order upon splitting off the glycan beyond theAsn78-bound GlcNAc. In particular, the Asn-bound GlcNAc shields the protein surface from the hydrophilicenvironment through interaction with predominantly hydrophobic amino acid residues located in bothtwisted -hairpins consisting of residues 10-28 and 59-84.
-subunit of human chorionicgonadotropin have been studied by NMR spectroscopy. Significant differences in chemical shifts andNOE intensities were observed for amino acid residues close to the carbohydrate chain at Asn78 upondeglycosylation beyond Asn78-bound GlcNAc. As no straightforward strategy is available for the calculationof the NMR structure of intact glycoproteins, a suitable computational protocol had to be developed. Tothis end, the X-PLOR carbohydrate force field designed for structure refinement was extended and modified.Furthermore, a computational strategy was devised to facilitate successful protein folding in the presenceof extended glycans during the simulation. The values for and dihedral angles of the glycosidiclinkages of the oligosaccharide core fragments GlcNAc2(1-4)GlcNAc1 and Man3(1-4)GlcNAc2 arerestricted to a limited range of the broad conformational energy minima accessible for free glycans. Thisdemonstrates that the protein core affects the dynamic behavior of the glycan at Asn78 by steric hindrance.Reciprocally, the NMR structures indicate that the glycan at Asn78 affects the stability of the proteincore. The backbone angular order parameters and displacement data of the generated conformers displayespecially for the -turn 20-23 a decreased structural order upon splitting off the glycan beyond theAsn78-bound GlcNAc. In particular, the Asn-bound GlcNAc shields the protein surface from the hydrophilicenvironment through interaction with predominantly hydrophobic amino acid residues located in bothtwisted -hairpins consisting of residues 10-28 and 59-84.