[20] Solid-phase protein hydration studies

Poole, P. L.; Finney, John

doi:10.1016/0076-6879(86)27023-8

Cited by 13 publications

(12 citation statements)

References 10 publications

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“…As mentioned previously, studies on lysozyme, subtilisin Carlsberg, α-chymotrypsin, and α-amylase show the onset of protein motions at ∼0.2 h. These have been correlated with the onset of enzyme catalysis in hydrated enzyme powders (23,38,86,102,109,110,119,140). However, one difficulty with measuring activity in enzyme powders of low hydration is the absence of a medium for the diffusion of substrate and product, particularly in enzymes such as lysozyme that have low specific activity.…”

Section: Effect Of Protein Hydration On Enzyme Activitymentioning

confidence: 87%

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The Role of Dynamics in Enzyme Activity

Daniel

Dunn

Finney

et al. 2003

Annu. Rev. Biophys. Biomol. Struct.

325

340

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Although protein function is thought to depend on flexibility, precisely how the dynamics of the molecule and its environment contribute to catalytic mechanisms is unclear. We review experimental and computational work relating to enzyme dynamics and function, including the role of solvent. The evidence suggests that fast motions on the 100 ps timescale, and any motions coupled to these, are not required for enzyme function. Proteins where the function is electron transfer, proton tunneling, or ligand binding may have different dynamical dependencies from those for enzymes, and enzymes with large turnover numbers may have different dynamical dependencies from those that turn over more slowly. The timescale differences between the fastest anharmonic fluctuations and the barrier-crossing rate point to the need to develop methods to resolve the range of motions present in enzymes on different time- and lengthscales.

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Section: Effect Of Protein Hydration On Enzyme Activitymentioning

confidence: 87%

“…A variety of observations made as enzymes are rehydrated from the anhydrous state to the solution (23, 25, 46, 102,109,110,112,140) have associated hydration changes with changes in protein-water interactions and protein dynamics. Based mainly on detailed work on lysozyme, the process seems to involve four stages.…”

Section: Structural Effects Of Solvationmentioning

confidence: 98%

The Role of Dynamics in Enzyme Activity

Daniel

Dunn

Finney

et al. 2003

Annu. Rev. Biophys. Biomol. Struct.

325

340

View full text Add to dashboard Cite

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“…Varying degrees of hydration were obtained by equilibrating preweighted amounts of HSA under a controlled humidity atmosphere, on a seven days period. Standard saturated salt solutions placed in a set of desiccators allowed one to obtain the atmosphere with controlled humidity [26]. Afterwards the powders were weighed again and the degree of hydration was determined from the weight difference; the water content of the lyophilized powders was estimated according to Schinkel et al [17] and used to correct the value of the hydration degree.…”

Section: Sample Preparationmentioning

confidence: 99%

Hydration and protein dynamics: an ESR and ST-ESR spin labelling study of human serum albumin

Marzola

Cannistraro

1992

Appl. Magn. Reson.

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Human serum albumin has been studied at low hydration level by the ESR spin labelling technique, under the assumption that a covalently bound spin-label is a reporter of the protein internal dynamics. At room temperature, the presence of a double component signal allowed us to monitor the influence of increasing hydration level on internal protein dynamics as well as on the superficial water dynamics. The ESR results have shown that the first 20 g of water per 100 g of protein activate the internal protein dynamics and that superficial water dynamics starts at higher hydration values. ESR expe¡ at low temperature have shown that at --l£176 ~ T~ -80~ the label experiences an increasing environmental polarity with increasing temperature in the samples with hydration values higher than about 20 g of water per 100 g of protein.The results are discussed in connection with both conformational substates of the protein and hydration water dynamics.

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“…20 μL (∼20 μg) of the diluted sample was subsequently placed on a high resistivity silicon window and excess water was removed by applying a low, steady flow of N 2 gas over the sample droplet for approximately 10 minutes. The resulting protein film was subsequently rehydrated by equilibrating the partially dried sample in a vacuum sealed container with the vapor pressure of a saturated salt solution at 20 • C for a minimum of 5 days [12]. For the experiments discussed in this manuscript, a relative humidity (RH) of 97% was obtained from the vapor pressure of a saturated K 2 SO4 solution [13].…”

Section: Sample Preparationmentioning

confidence: 99%

THz time scale structural rearrangements and binding modes in lysozyme-ligand interactions

Woods

2014

J Biol Phys

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Predicting the conformational changes in proteins that are relevant for substrate binding is an ongoing challenge in the aim of elucidating the functional states of proteins. The motions that are induced by protein-ligand interactions are governed by the protein global modes. Our measurements indicate that the detected changes in the global backbone motion of the enzyme upon binding reflect a shift from the large-scale collective dominant mode in the unbound state towards a functional twisting deformation that assists in closing the binding cleft. Correlated motion in lysozyme has been implicated in enzyme function in previous studies, but detailed characterization of the internal fluctuations that enable the protein to explore the ensemble of conformations that ultimately foster largescale conformational change is yet unknown. For this reason, we use THz spectroscopy to investigate the picosecond time scale binding modes and collective structural rearrangements that take place in hen egg white lysozyme (HEWL) when bound by the inhibitor (NAG) 3 . These protein thermal motions correspond to fluctuations that have a role in both selecting and sampling from the available protein intrinsic conformations that communicate function. Hence, investigation of these fast, collective modes may provide knowledge about the mechanism leading to the preferred binding process in HEWL-(NAG) 3 . Specifically, in this work we find that the picosecond time scale hydrogen-bonding rearrangements taking place in the protein hydration shell with binding modify the packing density within the hydrophobic core on a local level. These localized, intramolecular contact variations within the protein core appear to facilitate the large cooperative movements within the interfacial region separating the α-and β-domain that mediate binding. The THz time-scale fluctuations identified in the protein-ligand system may also reveal a molecular mechanism for substrate recognition.

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[20] Solid-phase protein hydration studies

Cited by 13 publications

References 10 publications

The Role of Dynamics in Enzyme Activity

The Role of Dynamics in Enzyme Activity

Hydration and protein dynamics: an ESR and ST-ESR spin labelling study of human serum albumin

THz time scale structural rearrangements and binding modes in lysozyme-ligand interactions

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