Ligand Binding Mechanics of Maltose Binding Protein

Morten Bertz, Matthias Rief

Research output: Contribution to journalArticle

33 Citations (Scopus)

Abstract

In the past decade, single-molecule force spectroscopy has provided new insights into the key interactions stabilizing folded proteins. A few recent studies probing the effects of ligand binding on mechanical protein stability have come to quite different conclusions. While some proteins seem to be stabilized considerably by a bound ligand, others appear to be unaffected. Since force acts as a vector in space, it is conceivable that mechanical stabilization by ligand binding is dependent on the direction of force application. In this study, we vary the direction of the force to investigate the effect of ligand binding on the stability of maltose binding protein (MBP). MBP consists of two lobes connected by a hinge region that move from an open to a closed conformation when the ligand maltose binds. Previous mechanical experiments, where load was applied to the N and C termini, have demonstrated that MBP is built up of four building blocks (unfoldons) that sequentially detach from the folded structure. In this study, we design the pulling direction so that force application moves the two MBP lobes apart along the hinge axis. Mechanical unfolding in this geometry proceeds via an intermediate state whose boundaries coincide with previously reported MBP unfoldons. We find that in contrast to N-C-terminal pulling experiments, the mechanical stability of MBP is increased by ligand binding when load is applied to the two lobes and force breaks the protein-ligand interactions directly. Contour length measurements indicate that MBP is forced into an open conformation before unfolding even if ligand is bound. Using mutagenesis experiments, we demonstrate that the mechanical stabilization effect is due to only a few key interactions of the protein with its ligand. This work illustrates how varying the direction of the applied force allows revealing important details about the ligand binding mechanics of a large protein.

Original languageEnglish
Pages (from-to)1097-1105
Number of pages9
JournalJournal of Molecular Biology
Volume393
Issue number5
DOIs
Publication statusPublished - 2009 Nov 13
Externally publishedYes

Fingerprint

Maltose-Binding Proteins
Mechanics
Ligands
Proteins
Maltose
Protein Stability
Mutagenesis

Keywords

  • atomic force microscopy
  • ligand binding
  • protein folding
  • single-molecule force spectroscopy
  • unfolding intermediate

ASJC Scopus subject areas

  • Molecular Biology

Cite this

Ligand Binding Mechanics of Maltose Binding Protein. / Bertz, Morten; Rief, Matthias.

In: Journal of Molecular Biology, Vol. 393, No. 5, 13.11.2009, p. 1097-1105.

Research output: Contribution to journalArticle

Bertz, Morten ; Rief, Matthias. / Ligand Binding Mechanics of Maltose Binding Protein. In: Journal of Molecular Biology. 2009 ; Vol. 393, No. 5. pp. 1097-1105.
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