Trehalose Prevents Myoglobin Collapse and Preserves Its Internal Mobility

Sastry, G. Madhavi; Agmon, Noam

doi:10.1021/bi9626057

Cited by 81 publications

(69 citation statements)

References 56 publications

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“…This "upper limit" frequency is the same as that which we obtain from samples of COMb embedded in an ultra high viscosity glass matrix derived from trehalose. Earlier studies show that increasing the viscosity slows and eventually stops the tertiary relaxation of photodissociated COMb (23,25,28,59,60,62,64,(81)(82)(83)(84)(85)(86). These findings all indicate that the distribution of photoproduct conformations being probed for the [COMb] ϩ100% glycerol sample is reflective of the initial liganded protein but with the iron displaced out of the heme plane (and the heme geometry appropriately modified).…”

Section: Discussionmentioning

confidence: 80%

Spectroscopically and Kinetically Distinct Conformational Populations of Sol-Gel-encapsulated Carbonmonoxy Myoglobin

Samuni

Dantsker

Khan

et al. 2002

Journal of Biological Chemistry

108

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We have used sol-gel encapsulation protocols to trap kinetically and spectroscopically distinct conformational populations of native horse carbonmonoxy myoglobin. The method allows for direct comparison of functional and spectroscopic properties of equilibrium and non-equilibrium populations under the same temperature and viscosity conditions. The results implicate tertiary structure changes that include the proximal heme environment in the mechanism for population-specific differences in the observed rebinding kinetics. Differences in the resonance Raman frequency of (Fe-His), the iron-proximal histidine stretching mode, are attributed to differences in the positioning of the F helix. For myoglobin, the degree of separation between the F helix and the heme is assigned as the conformational coordinate that modulates both this frequency and the innermost barrier controlling CO rebinding. A comparison with the behavior of encapsulated derivatives of human adult hemoglobin indicates that these CO binding-induced conformational changes are qualitatively similar to the tertiary changes that occur within both the R and T quaternary states. Protein-specific differences in the time scale for the proposed F helix relaxation are attributed to variations in the intra-helical hydrogen bonding patterns that help stabilize the position of the F helix. Myoglobin (Mb)1 continues to be used as a model protein system for investigations into how structure, dynamics, and reactivity interconnect to give rise to functionality. Recently there has been a renewed interest in myoglobin based on several new developments. From a functional point of view there are indications and suggestions that, apart from its physiological importance in facilitating oxygen diffusion, myoglobin has functions arising from its ability to participate in catalytic multisubstrate reactions involving such molecules as NO, O 2 , and H 2 O 2 (1-3). In addition the presence of apolar intra-protein substrate docking sites (the so-called Xe cavities) (4, 5) have recently been linked both to these newly proposed functions and to the kinetic patterns associated with geminate and bimolecular ligand binding (3, 6 -13).These new developments highlight fundamental biophysical issues that have not been fully addressed despite the extensive research effort that has been directed toward myoglobin. The issue that we address in this work relates to ligand reactivity as a function of conformation. Several studies show that myoglobin can adopt different tertiary conformations. In particular there is evidence that the equilibrium distribution of tertiary conformations is different for the deoxy (ferrous five coordinate) and CO derivatives of Mb. X-ray crystallographic studies show that there is a small clam shell-like rotation of the E and F helices in going from deoxy to COMb (14) as well as adjustments in the positioning of residues in the distal heme pocket (8 -10, 15, 16) that may or may not be related to the clam shell motion. Recent time-resolved x-ray crystallographic stud...

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Section: Discussionmentioning

confidence: 80%

Spectroscopically and Kinetically Distinct Conformational Populations of Sol-Gel-encapsulated Carbonmonoxy Myoglobin

Samuni

Dantsker

Khan

et al. 2002

Journal of Biological Chemistry

108

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“…The importance of an evolving reaction coordinate as a factor in the control of rebinding rates in Mb was first put forth by Agmon and co-workers (73)(74)(75). Scott and Friedman (76) have shown this effect experimentally in hemoglobin where the fast tertiary relaxation does control geminate ligand rebinding at ambient temperatures.…”

Section: Factors Influencing Rebinding From Within the Distal Heme Pomentioning

confidence: 97%

The Position 68(E11) Side Chain in Myoglobin Regulates Ligand Capture, Bond Formation with Heme Iron, and Internal Movement into the Xenon Cavities

Dantsker

Roche

Samuni

et al. 2005

Journal of Biological Chemistry

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After photodissociation, ligand rebinding to myoglobin exhibits complex kinetic patterns associated with multiple first-order geminate recombination processes occurring within the protein and a simpler bimolecular phase representing second-order ligand rebinding from the solvent. A smooth transition from cryogeniclike to solution phase properties can be obtained by using a combination of sol-gel encapsulation, addition of glycerol as a bathing medium, and temperature tuning (؊15 3 65°C). This approach was applied to a series of double mutants, myoglobin CO (H64L/ V68X, where X ‫؍‬ Ala, Val, Leu, Asn, and Phe), which were designed to examine the contributions of the position 68(E11) side chain to the appearance and disappearance of internal rebinding phases in the absence of steric and polar interactions with the distal histidine. Based on the effects of viscosity, temperature, and the stereochemistry of the E11 side chain, the three major phases, B 3 A, C 3 A, and D 3 A, can be assigned, respectively, to ligand rebinding from the following: (i) the distal heme pocket, (ii) the xenon cavities prior to large amplitude side chain conformational relaxation, and (iii) the xenon cavities after significant conformational relaxation of the position 68(E11) side chain. The relative amplitudes of the B 3 A and C 3 A phases depend markedly on the size and shape of the E11 side chain, which regulates sterically both ligand return to the heme iron atom and ligand migration to the xenon cavities. The internal xenon cavities provide a transient docking site that allows side chain relaxations and the entry of water into the vacated distal pocket, which in turn slows ligand recombination markedly.Proteins are inherently complex materials, and even their simplest reactions exhibit layers of complexity that were not anticipated a few decades ago (1-3). A case in point is carbon monoxide (CO) binding to the heme iron atom in myoglobin (Mb) 3 (4). Much of the work on Mb has been directed toward understanding the biophysical principles associated with both bimolecular and internal ligand rebinding after photolysis of the Fe-CO bond. Key issues include the following: (i) the roles of distal and proximal heme pocket amino acids; (ii) the roles of internal water molecules near the active site; (iii) the roles of local and global conformational relaxations that are modulated by solvent; and (iv) the roles of pre-existing internal cavities associated with xenon binding. Time-resolved spectroscopic and x-ray crystallographic studies have demonstrated that dissociated ligands can access internal cavities that arise from packing defects in the globin tertiary structure (5-26). A remaining challenge is to establish quantitatively how all these factors contribute to the multiple kinetic phases associated with internal ligand binding at cryogenic temperatures and high viscosity and to those observed at ambient temperatures and physiologically relevant viscosities (21, 27-37).Much of the cryogenic work and the time-resolved x-ray crystallograp...

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“…FT-IR and other studies have pointed to the gradual replacement of water by sugars, mainly trehalose, but also polymers like polyethylene glycol, and so forth. [84][85][86][87] In many cases where comparisons have been carried out, trehalose has come out at the top, not only among disaccharides but in comparison with other excipients as well. 86,87 Simultaneous use of trehalose and ionic salts has been shown to have a synergistic effect on the freeze-drying stability of hen egg white lysozyme.…”

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confidence: 99%

“…[84][85][86][87] In many cases where comparisons have been carried out, trehalose has come out at the top, not only among disaccharides but in comparison with other excipients as well. 86,87 Simultaneous use of trehalose and ionic salts has been shown to have a synergistic effect on the freeze-drying stability of hen egg white lysozyme. 88 This stabilization results from the interaction of trehalose with the protein, as confirmed by changes in the CO stretching vibration, resulting from the destructuring of the surrounding water molecules.…”

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confidence: 99%

Effect of trehalose on protein structure

Jain¹,

Roy²

2008

Protein Science

714

576

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Trehalose is a ubiquitous molecule that occurs in lower and higher life forms but not in mammals. Till about 40 years ago, trehalose was visualized as a storage molecule, aiding the release of glucose for carrying out cellular functions. This perception has now changed dramatically. The role of trehalose has expanded, and this molecule has now been implicated in a variety of situations. Trehalose is synthesized as a stress-responsive factor when cells are exposed to environmental stresses like heat, cold, oxidation, desiccation, and so forth. When unicellular organisms are exposed to stress, they adapt by synthesizing huge amounts of trehalose, which helps them in retaining cellular integrity. This is thought to occur by prevention of denaturation of proteins by trehalose, which would otherwise degrade under stress. This explanation may be rational, since recently, trehalose has been shown to slow down the rate of polyglutamine-mediated protein aggregation and the resultant pathogenesis by stabilizing an aggregation-prone model protein. In recent years, trehalose has also proved useful in the cryopreservation of sperm and stem cells and in the development of a highly reliable organ preservation solution. This review aims to highlight the changing perception of the role of trehalose over the last 10 years and to propose common mechanisms that may be involved in all the myriad ways in which trehalose stabilizes protein structures. These will take into account the structure of trehalose molecule and its interactions with its environment, and the explanations will focus on the role of trehalose in preventing protein denaturation.

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Trehalose Prevents Myoglobin Collapse and Preserves Its Internal Mobility

Cited by 81 publications

References 56 publications

Spectroscopically and Kinetically Distinct Conformational Populations of Sol-Gel-encapsulated Carbonmonoxy Myoglobin

Spectroscopically and Kinetically Distinct Conformational Populations of Sol-Gel-encapsulated Carbonmonoxy Myoglobin

The Position 68(E11) Side Chain in Myoglobin Regulates Ligand Capture, Bond Formation with Heme Iron, and Internal Movement into the Xenon Cavities

Effect of trehalose on protein structure

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