TβR-II discriminates the high- and low-affinity TGF-β isoforms via two hydrogen-bonded ion Pairs

Jason Baardsnes, Cynthia S. Hinck, Andrew P. Hinck, Maureen D. O'Connor-McCourt

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33 Citations (Scopus)

Abstract

The TGF-β isoforms, TGF-β1, -β2, and-β3, share greater than 70% sequence identity and are almost structurally identical. TGF-β2 differs from the others, however, in that it binds the TGF-β type II receptor (TβR-II) with much lower affinity than either TGF-β1 or-β3. It has been previously shown that three conserved interfacial residues, Arg25, Val92, Arg94, in TGF-β1 and-β3 are responsible for their high-affinity interaction with TβR-II. In this study, the role of each of these residues was examined by creating single, double, and triple substitutions resulting in both TGF-β3 loss-of-function and TGF-β2 gain-of-function variants. One-dimensional 1H NMR spectra of the variants confirmed a lack of large structural perturbations. Affinities, kinetics, and thermodynamics for TβR-II binding were determined by surface plasmon resonance biosensor analysis. Double substitutions revealed that nearly all of the high-affinity binding is contributed by Arg25 and Arg94. Single site substitutions showed that Arg94 makes the greatest contribution. Substitution of Arg25 and Arg94 with alanine verified the requirement of the arginine guanidinium functional groups for the highly specific hydrogen-bonded ion pairs formed between Arg25 and Arg94 of TGF-β1 and-β3, and Glu119 and Asp32 of TβR-II. Further kinetic and thermodynamic analyses confirmed that Arg25 and Arg94 are primarily responsible for high-affinity binding and also revealed that noninterfacial longer range effects emanating from the TGF-β structural framework contribute slightly to TβR-II binding. Growth inhibition assays showed that binding changes generally correlate directly with changes in function; however, a role Val92 in this cellular context was uncovered.

Original languageEnglish (US)
Pages (from-to)2146-2155
Number of pages10
JournalBiochemistry
Volume48
Issue number10
DOIs
StatePublished - Mar 17 2009

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Thermodynamics
Protons
Hydrogen
Protein Isoforms
Substitution reactions
Ions
Surface Plasmon Resonance
Guanidine
Biosensing Techniques
Alanine
Arginine
Kinetics
Surface plasmon resonance
Growth
Biosensors
Functional groups
Assays
Nuclear magnetic resonance
Proton Magnetic Resonance Spectroscopy

ASJC Scopus subject areas

  • Biochemistry

Cite this

Baardsnes, J., Hinck, C. S., Hinck, A. P., & O'Connor-McCourt, M. D. (2009). TβR-II discriminates the high- and low-affinity TGF-β isoforms via two hydrogen-bonded ion Pairs. Biochemistry, 48(10), 2146-2155. https://doi.org/10.1021/bi8019004

TβR-II discriminates the high- and low-affinity TGF-β isoforms via two hydrogen-bonded ion Pairs. / Baardsnes, Jason; Hinck, Cynthia S.; Hinck, Andrew P.; O'Connor-McCourt, Maureen D.

In: Biochemistry, Vol. 48, No. 10, 17.03.2009, p. 2146-2155.

Research output: Contribution to journalArticle

Baardsnes, J, Hinck, CS, Hinck, AP & O'Connor-McCourt, MD 2009, 'TβR-II discriminates the high- and low-affinity TGF-β isoforms via two hydrogen-bonded ion Pairs', Biochemistry, vol. 48, no. 10, pp. 2146-2155. https://doi.org/10.1021/bi8019004
Baardsnes, Jason ; Hinck, Cynthia S. ; Hinck, Andrew P. ; O'Connor-McCourt, Maureen D. / TβR-II discriminates the high- and low-affinity TGF-β isoforms via two hydrogen-bonded ion Pairs. In: Biochemistry. 2009 ; Vol. 48, No. 10. pp. 2146-2155.
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