Nuclear magnetic resonance structural and ligand binding studies of BLBC, a two-domain fragment of barley lectin

Biochemistry
J L Weaver, James H Prestegard

Abstract

Plant lectins are useful targets for biophysical studies of protein-carbohydrate recognition, a process of general interest because of its many roles in human physiology. Here, nuclear magnetic resonance (NMR) based structural and carbohydrate binding data on a two-domain fragment of the normally four-domain barley lectin protein are presented. The structural data, while preliminary, clearly shows that the recombinantly produced simplified model system, called BLBC, retains a nativelike fold. However, unlike the full-length parent protein, which is dimeric, BLBC is shown by pulsed-field gradient NMR diffusion studies to be largely monomeric. Still, the fragment retains nativelike carbohydrate binding properties. These properties are examined in some detail using heteronuclear single quantum coherence (HSQC) NMR spectroscopy on a uniformly 15N-labeled sample. Ligand-induced chemical shift changes in the 1H-15N HSQC spectrum are monitored as 15N-labeled BLBC is titrated with increasing concentrations of the unlabeled carbohydrate, N,N',N"-triacetylchitotriose. Well-resolved resonances from the individual domains show that BLBC binds ligand at two distinct and independent ligand binding sites, one in each domain. Binding constants...Continue Reading

References

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Citations

Apr 26, 2001·NMR in Biomedicine·K NicolayM J Kruiskamp
Jul 17, 1998·Nature Structural Biology·J H Prestegard
Oct 30, 2010·Nature Protocols·Molly J RossowEnrico Gratton
Apr 25, 2013·Progress in Nuclear Magnetic Resonance Spectroscopy·Quentin Kaas, David J Craik
Oct 12, 2004·Insect Biochemistry and Molecular Biology·Toru TogawaSusumu Izumi
May 26, 1999·Journal of Magnetic Resonance·J A LosoncziJ H Prestegard
Apr 20, 2000·The Journal of Biological Chemistry·T SuetakeK Kawano
Dec 2, 2004·Molecular and Cellular Biology·Séverine Celton-MorizurAnne Paoletti
Jul 18, 2001·Journal of the American Chemical Society·J C HusM Blackledge
Jun 21, 2001·Journal of the American Chemical Society·J MeilerR Brüschweiler

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