Kinetics of breaking a salt-bridge critical in protein unfolding

Gruia, Andreea D.; Fischer, Stefan; Smith, Jeremy C.

doi:10.1016/j.cplett.2003.12.038

Cited by 15 publications

(15 citation statements)

References 34 publications

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“…In either case, two strong interactions dominated by a buried salt bridge can form between the Asp residue of MUC1 and the Ab. Potential mean force calculations for similar salt bridges show bond widths that range from 2.2 to 3.0 Å (47,48), which compare well with the characteristic rupture distance of 2.8 Å measured in the force spectroscopy experiments.…”

Section: Muc1 Docking Simulationssupporting

confidence: 64%

Dynamic force spectroscopy of parallel individual Mucin1–antibody bonds

Sulchek

Friddle

Langry

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

165

215

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We used atomic force microscopy to measure the binding forces between Mucin1 (MUC1) peptide and a single-chain variable fragment (scFv) antibody selected from a scFv library screened against MUC1. This binding interaction is central to the design of molecules used for targeted delivery of radioimmunotherapeutic agents for prostate and breast cancer treatment. Our experiments separated the specific binding interaction from nonspecific interactions by tethering the antibody and MUC1 molecules to the atomic force microscope tip and sample surface with flexible polymer spacers. Rupture force magnitude and elastic characteristics of the spacers allowed identification of the rupture events corresponding to different numbers of interacting proteins. We used dynamic force spectroscopy to estimate the intermolecular potential widths and equivalent thermodynamic off rates for monovalent, bivalent, and trivalent interactions. Measured interaction potential parameters agree with the results of molecular docking simulation. Our results demonstrate that an increase of the interaction valency leads to a precipitous decline in the dissociation rate. Binding forces measured for monovalent and multivalent interactions match the predictions of a Markovian model for the strength of multiple uncorrelated bonds in a parallel configuration. Our approach is promising for comparison of the specific effects of molecular modifications as well as for determination of the best configuration of antibody-based multivalent targeting agents.atomic force microscopy ͉ multivalency ͉ radioimmunmotherapy ͉ binding affinity I nteractions between biological molecules drive a vast variety of cellular processes and span a wide range of strength and complexity. Multivalent interactions where several binding units combine to produce superior binding strength play an important role in adaptive immune response (1) and intercellular adhesion (2), as well as in the mechanism of action of many pharmaceuticals (3). Clinical researchers have used multivalency as an affinity-enhancing approach (4, 5) in a variety of immunotherapies and imaging techniques to target specific tissues (6, 7).Linking several molecules into a large multivalent binding construct also creates bulky agents that exhibit reduced tissue penetration and have a higher probability of accumulation in liver (8). Therefore, a better understanding of the multivalent binding is necessary for the creation of optimized agents that balance binding efficiency and molecular size. Quantitative characterization of multivalent interactions is also important for understanding the basic biophysics of complex molecular systems.The last decade saw an explosion of interaction force measurement techniques that allowed researchers to measure and apply molecular level stresses (9-11). Atomic force microscopy (AFM) probes ligand-receptor interactions by simply pulling off the ligand from the receptor using external force (12). Kinetic approaches to the binding force measurements, such as dynamic force spectroscopy ...

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Section: Muc1 Docking Simulationssupporting

confidence: 64%

Dynamic force spectroscopy of parallel individual Mucin1–antibody bonds

Sulchek

Friddle

Langry

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

165

215

View full text Add to dashboard Cite

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“…The breaking and forming of salt bridges is estimated to occur in nanoseconds making them ideal for participating in this process (27,28). Here, we provide evidence that a salt bridge between ␤E153 and ␤K196 located on ␤7 and ␤9 of the GABA A R is important for regulating loop C movement.…”

Section: Discussionmentioning

confidence: 69%

A conserved salt bridge critical for GABA _A receptor function and loop C dynamics

Venkatachalan

Czajkowski²

2008

Proc. Natl. Acad. Sci. U.S.A.

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The authors note that in Fig. 1 the alignment of the nicotinic receptor ␣1 subunit sequences (Torpedo californica and rat) in the loop C region with the GABA A receptor ␤ subunit sequences aligns Y190 and T191 of the former with K196 and K197 of the latter. The alignment should align H186 and W197 of the nicotinic receptor with K196 and K197 of the GABA A receptor. The nicotinic sequence that reads VYYTCCPD 195 should read WKHW..YT 191. The error does not alter the conclusions of this article. The corrected figure and its legend appear below. MEDICAL SCIENCESCorrection for ''Abnormal integrin-mediated regulation of chronic myelogenous leukemia CD34 ϩ cell proliferation: BCR/ ABL up-regulates the cyclin-dependent kinase inhibitor, p27 Kip , which is relocated to the cell cytoplasm and incapable of regulating cdk2 activity,''

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“…Estimates made by MD simulation have shown that salt bridges can exist unbroken substantially longer than 1 ns, even as long as 200 ns (42,84,85). A representative example can be given with the case of the Braun's E. coli outer membrane lipoprotein protein (Lpp-56).…”

Section: Do Proteins Benefit From Salt Bridges?mentioning

confidence: 99%

Defining the Role of Salt Bridges in Protein Stability

Jelesarov

Karshikoff

2008

Methods in Molecular Biology

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Running head: Salt bridges and protein stability 2 (i) SummaryAlthough the energetic balance of forces stabilizing proteins has been established qualitatively over the last decades, quantification of the energetic contribution of particular interactions still poses serious problems. The reasons are the strong cooperativity and the interdependence of noncovalent interactions. Salt bridges are a typical example. One expects that ionizable side chains frequently form ion pairs in innumerable crystal structures. Since electrostatic attraction between opposite charges is strong per se, salt bridges can intuitively be regarded as an important factor stabilizing the native structure. Is that really so? In this chapter we critically reassess the available methods to delineate the role of electrostatic interactions and salt bridges to protein stability, and discuss the progress and the obstacles in this endeavor. The basic problem is that formation of salt bridges depends on the ionization properties of the participating groups, which is significantly influenced by the protein environment. Furthermore, salt bridges experience thermal fluctuations, continuously break and re-form, and their lifespan in solution is governed by the flexibility of the protein. Finally, electrostatic interactions are long-range and might be significant in the unfolded state, thus seriously influencing the energetic profile. Elimination of salt bridges by protonation/deprotonation at extreme pH or by mutation provides only rough energetic estimates, since there is no way to account for the non-additive response of the protein moiety. From what we know so far, the strength of electrostatic interactions is strongly contextdependent, yet it is unlikely that salt bridges are dominant factors governing protein stability.Nevertheless, proteins from thermophiles and hyperthermophiles exhibit more, and frequently networked, salt bridges than proteins from the mesophilic counterparts. Increasing the thermal (not the thermodynamic) stability of proteins by optimization of charge-charge interactions is a good example for an evolutionary solution utilizing physical factors.(ii) Keywords: electrostatic interactions, salt bridge, protein stability, thermal stability, denatured state, pK, protein unfolding 3

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Kinetics of breaking a salt-bridge critical in protein unfolding

Cited by 15 publications

References 34 publications

Dynamic force spectroscopy of parallel individual Mucin1–antibody bonds

Dynamic force spectroscopy of parallel individual Mucin1–antibody bonds

A conserved salt bridge critical for GABA _A receptor function and loop C dynamics

Defining the Role of Salt Bridges in Protein Stability

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Kinetics of breaking a salt-bridge critical in protein unfolding

Cited by 15 publications

References 34 publications

Dynamic force spectroscopy of parallel individual Mucin1–antibody bonds

Dynamic force spectroscopy of parallel individual Mucin1–antibody bonds

A conserved salt bridge critical for GABA A receptor function and loop C dynamics

Defining the Role of Salt Bridges in Protein Stability

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A conserved salt bridge critical for GABA _A receptor function and loop C dynamics