Water at the Macromolecular Surface

Parsegian, V. Adrian; Rand, R. P.; Colombo, Marcio F.; Rau, Donald C.

doi:10.1021/ba-1994-0235.ch009

Cited by 13 publications

(14 citation statements)

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“…By using two different model approaches, the Wyman linkage equation or the Gibbs Duhem equation, we found that 65-72 1 water molecules are linked to the binding of four oxygen molecules to human Hb under the experimental conditions used. Furthermore, this hydration change was found to be in agreement with the difference between the solvent-accessible surface areas of deoxy-and oxy-Hb computed from x-ray structures by Clothia et al (5,6).The experimental approach and analyses originally used by us to assess and quantify the role of water on the allosteric control of Hb have been tested in several other biochemical systems (7)(8)(9)(10)(11)(12)(13)(14). A standard linear dependence of equilibrium and kinetic free energy parameters on water activity rather than solute activity has been lately reported.…”

supporting

confidence: 78%

“…The experimental approach and analyses originally used by us to assess and quantify the role of water on the allosteric control of Hb have been tested in several other biochemical systems (7)(8)(9)(10)(11)(12)(13)(14). A standard linear dependence of equilibrium and kinetic free energy parameters on water activity rather than solute activity has been lately reported.…”

mentioning

confidence: 99%

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The Water Effect on Allosteric Regulation of Hemoglobin Probed in Water/Glucose and Water/Glycine Solutions

Colombo

Bonilla-Rodriguez

1996

Journal of Biological Chemistry

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We present data which indicate that glycine and glucose influence HbA oxygen affinity to the same extent, despite the fact that glycine increases and glucose decreases the bulk dielectric constant of the solution. Furthermore, we derive an equation linking changes in oxygen affinity to changes in differential solute and water binding to test critically the possibility of neutral solute heterotropic binding. Applied to the data, these analyses support our original interpretation that neutral solutes act indirectly on the regulation of allosteric behavior of hemoglobin by varying the chemical potential of water in solution. This leads to a displacement of the equilibrium between Hb conformational states in proportion to their differential hydration.It is well established that differential water binding occurs between distinct conformations of a protein. Hence, whether or not differential water binding contributes to the energetic of protein reactions, as some ions and metabolites do, is a key question to fully comprehend the mechanism of biological control. The role of water on allosteric regulation had been previously investigated, using Hb oxygenation as a model, by the so-called osmotic stress method (1, 2). The concept of this method is to use solutes, which are potentially excluded from the protein surface, to set the bulk water activity a w (3). Haire and Hedlund (4), using ethylene glycol, observed that at a low concentration range ethylene glycol decreases Hb oxygen affinity while at a higher concentration causes the opposite effect. They interpreted these results as reflecting preferential binding of ethylene glycol to Hb at the low solute concentration range, whereas, at high ethylene glycol concentration range, the increased O 2 binding affinity of Hb was attributed to an effect of water activity on the R to T allosteric equilibrium (4). In our former studies of the effect of water activity on Hb O 2 binding properties, we have used chemically different solutes like sucrose, stachyose, and polyethylene glycols to distinguish solute from water binding to Hb (1). Besides finding evidences for an indirect effect of solutes through water on the deoxy-to oxy-Hb conformational equilibrium, we have also determined quantitatively this solvation effect on Hb allosteric transition in terms of the differential number of water molecules bound between the two extreme conformations of Hb. By using two different model approaches, the Wyman linkage equation or the Gibbs Duhem equation, we found that 65-72 1 water molecules are linked to the binding of four oxygen molecules to human Hb under the experimental conditions used. Furthermore, this hydration change was found to be in agreement with the difference between the solvent-accessible surface areas of deoxy-and oxy-Hb computed from x-ray structures by Clothia et al. (5,6).The experimental approach and analyses originally used by us to assess and quantify the role of water on the allosteric control of Hb have been tested in several other biochemical systems (7)(8)(9)...

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supporting

confidence: 78%

mentioning

confidence: 99%

The Water Effect on Allosteric Regulation of Hemoglobin Probed in Water/Glucose and Water/Glycine Solutions

Colombo

Bonilla-Rodriguez

1996

Journal of Biological Chemistry

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“…In cases where the free and ligand-bound forms of a protein are differentially solvated, osmotic pressure will shift the binding equilibrium toward the protein conformation that is in the less hydrated form (15)(16)(17)(18). For osmotic pressure studies, PEG-8000 is an optimal "nonbinding" high molecular weight cosolute compared with either sucrose or glycerol on several accounts (15-18).…”

Section: Reaction Step Rate Constant Valuementioning

confidence: 99%

“…Protecting osmolytes are routinely added to enzyme preparations to increase enzyme stability during freezing and storage. The emerging view is that protecting osmolytes raise the free energy of a more hydrated partially "unfolded" state, which favors a less hydrated and more "folded" population (15)(16)(17)(18). Accordingly, we may postulate that nucleotide clamping results in some peptide region(s) becoming more unfolded and solvent exposed.…”

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confidence: 99%

Kinetic Mechanism of Fully Activated S6K1 Protein Kinase

Keshwani

Harris

2008

Journal of Biological Chemistry

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S6K1 is a member of the AGC subfamily of serine-threonine protein kinases, whereby catalytic activation requires dual phosphorylation of critical residues in the conserved T-loop (Thr-229) and hydrophobic motif (Thr-389). Previously, we described production of the fully activated catalytic kinase domain construct, His 6 -S6K1␣II(⌬AID)-T389E. Now, we report its kinetic mechanism for catalyzing phosphorylation of a model peptide substrate (Tide, RRRLSSLRA A key requirement for higher eukaryotic cells in sustaining prolific capacity is growth regulation, whereby increasing cellular mass and size prerequisite to division derive from coordinate macromolecular biosynthesis. The 70-kDa 40 S ribosomal protein S6 kinase-1 (S6K1) 2 is a key enzyme in coordinating cell growth with proliferation, as mitogen, nutrient, and energy status signaling pathways converge to activate S6K1 and initiate protein translation (1-5). Two S6K1 isoforms (accession no. NM003161, ␣I and ␣II isoforms) are produced from a single gene by alternative mRNA splicing and the use of an alternative translational start site (6). The 525-residue ␣I isoform contains an N-terminal 23 residue segment that encodes a polybasic nuclear localization motif, whereas the cytoplasmic ␣II isoform starts at a Met residue equivalent to Met-24 in the ␣I isoform, and the sequences of both isoforms are identical thereafter.S6K1 is a member of the AGC subfamily of serine-threonine protein kinases in which amino acid sequences are conserved in a segment of the catalytic kinase domain known as the activation loop or T-loop, as well as in a segment near the C terminus of the kinase domain known as the hydrophobic motif (7). Similar to other AGC kinase family members, catalytic activation of S6K1 minimally requires dual phosphorylation of a critical residue in both the T-loop and hydrophobic motif. For the fulllength S6K1␣I isoform these residues correspond to Thr-252 and Thr-412, respectively (8), whereas in the S6K1␣II isoform the identical residues correspond to Thr-229 and Thr-389 (9). With combined knowledge from available amino acid sequence alignments and x-ray structures, molecular modeling and biochemical testing now provide strong evidence for a common AGC kinase activation mechanism in which the C-terminal phosphorylated hydrophobic motif interacts with a phosphate binding pocket located in the small N-lobe of the kinase (10). This intramolecular interaction acts synergistically with T-loop phosphorylation to stabilize the active conformation, whereby a critical Glu residue in the ␣C-helix forms an ion pair with the catalytic Lys that functions to position the terminal phosphate of ATP for phosphotransfer in the kinase reaction.In recognizing the synergistic role of AGC kinase dual site phosphorylation, neither an x-ray three-dimensional structure nor a kinetic mechanism has been reported for any S6K1 isoform or domain construct. This derives largely from the inability to generate fully Thr-229 phosphorylated and activated S6K1. In previous work, we demonstrated b...

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“…Most data were fitted to DVLO theory with constant and dissimilar surface potential boundary conditions (20) and nonretarded van der Waals interaction (26). However, for hydrophobic interactions an additional acid-base (AB) attractive term was added (27), and the final expression for force normalized with the particle radius (F/R) in N/m is [8] where ε 0 is the permittivity of free space (C 2 /J m), κ is the reciprocal Debye length (m Interactions in deionized water. Results for experiments conducted in deionized water (pH ∼ 6) are presented graphically in Fig.…”

Section: Effect Of Surface Hydrophobicitymentioning

confidence: 99%