Thermal transitions in the purple membrane from <i>Halobacterium halobium</i>

Shnyrov, Valery L.; Mateo, Pedro L.

doi:10.1016/0014-5793(93)81400-t

Cited by 49 publications

(47 citation statements)

References 24 publications

(12 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…8O"C, attributed to a structural change in the crystal lattice of the membrane, followed by an irreversible transition at ca. lOO"C, associated with partial thermal unfolding of the membrane protein [16][17][18][19][20][21][22][23]. In the present study the thermal behavior of partially delipidated PM as a function of pH and bound cations removal has been compared to that of native PM.…”

Section: Iutroductionmentioning

confidence: 99%

Thermal stability of lipid‐depleted purple membranes at neutral and low pH values

1994

View full text Add to dashboard Cite

Differential scanning calorimetry was used to compare the thermal behavior of native and delipidated purple membrane fragments at pH values corresponding to purple, blue and acid-purple forms. At neutral pH, delipidation results in a 2.5-to 3-times increase in the cooperativity of the denaturational transition, accompanied by a minor increase in its temperature. At pH values below 5 the delipidated membranes exhibit considerably higher thermal stability than the native membranes. The reversible predenaturational transition observed in the native state is not detectable upon delipidation. There is no strict correlation between color changes upon acid&cation and deionization of either native or delipidated purple membranes and their thermal stability.

show abstract

Section: Iutroductionmentioning

confidence: 99%

Thermal stability of lipid‐depleted purple membranes at neutral and low pH values

1994

View full text Add to dashboard Cite

show abstract

“…This would be a reasonable possibility for AChE, which is a large protein and could, in principle, be composed of several domains (Privalov, 1982;Garel, 1992). We analyzed this possibility by application of successive annealing procedures (Shnyrov et al, 1984;Shnyrov & Mateo, 1993;Shnyrov, 1994). Thus, AChE was first heated in the microcalorimeter cell to a temperature of 42 "C, which would be close to the maximum for a putative first transition.…”

mentioning

confidence: 99%

Irreversible thermal denaturation ofTorpedo californicaacetylcholinesterase

et al. 1995

Self Cite

View full text Add to dashboard Cite

Thermal denaturation of Torpedo culifornicu acetylcholinesterase, a disulfide-linked homodimer with 537 amino acids in each subunit, was studied by differential scanning calorimetry. It displays a single calorimetric peak that is completely irreversible, the shape and temperature maximum depending on the scan rate. Thus, thermal denaturation of acetylcholinesterase is an irreversible process, under kinetic control, which is described well by the twostate kinetic scheme N 5 D, with activation energy 131 * 8 kcal/mol. Analysis of the kinetics of denaturation in the thermal transition temperature range, by monitoring loss of enzymic activity, yields activation energy of 121 t 20 kcal/mol, similar to the value obtained by differential scanning calorimetry. Thermally denatured acetylcholinesterase displays spectroscopic characteristics typical of a molten globule state, similar to those of partially unfolded enzyme obtained by modification with thiol-specific reagents. Evidence is presented that the partially unfolded states produced by the two different treatments are thermodynamically favored relative to the native state.Keywords: acetylcholinesterase; differential scanning calorimetry; irreversible denaturation; molten globule; thioldisulfide exchange; two-state kinetic model Folding of many small globular proteins is a cooperative process in the course of which only two states, the fully unfolded state, U, and the native state, N, are significantly populated. In the case of large proteins, which are generally believed to contain several domains (Jaenicke, 1991;Garel, 1992), folding has generally been considered to be cooperative within each domain, the only species populated in the course of either folding or unfolding being combinations of completely folded and completely unfolded domains (Privalov, 1982;Brandts et al., 1989;Garel, 1992). Evidence is accumulating, however, that another state, intermediate between N and U, can exist. This is a compact state that lacks the unique tertiary structure of the native protein but possesses substantial secondary structure. This state has been named the molten globule (MG) state (Kuwajima, 1989;Kim & Baldwin, 1990;Ptitsyn, 1992). The MG state is considered to serve as an intermediate on the pathway from the nascent polypeptide chain to the fully folded native protein in vivo (Gething & Sambrook, 1992). For most proteins, the MG state is unstable under physiological conditions and readily converts to the N state in vitro (Ptitsyn, 1992 Because the definition of the MG state is controversial (see, for example, Griko et al., 1994; Ewbank et al., 1995;Okazaki et al., 1995), in the present paper this term is used to refer to compact states of acetylcholinesterase (AChE) that preserve substantial secondary structure and lack most of the tertiary structure of the native enzyme. We have shown recently that exposure of a native dimeric form of Torpedo AChE to various treatments generates long-lived partially unfolded species displaying many of the characteristics of the MG...

show abstract

“…Upon heating the PM sample, the molecules become more labile and the residual quadrupolar splitting, which seems to reflect the organization of the molecules in the PM microcrystal, decreased significantly. If heating is performed outside the magnet, the process is reversible, as suggested from calorimetric studies; 44 namely the residual quadrupolar splitting of the deuterium resonance of the D 2 O molecules is observed following cooling of the sample to 298 K. However, a similar heating and cooling process performed in a strong magnetic field results in a somewhat different organization in which the residual quadrupolar splitting of the D 2 O is not observed. Therefore, the process appears to be irreversible.…”

Section: Discussionmentioning

confidence: 91%

“…44 According to these studies, the transitions, which occur below 343 K, are reversible. Only the three transitions that occur above this temperature are considered irreversible.…”

Section: The Effect Of the Magnetic Fieldmentioning

confidence: 97%

“…Only the three transitions that occur above this temperature are considered irreversible. 44 Therefore, we suspected that the magnetic field has an additional effect on the arrangement and reorganization of the system, which in turn affects the NMR characteristics of the water molecules. Indeed, when the sample was heated to 338 K outside the magnet, the process was reversible and similar doublets were observed in the 2 H single-pulse spectrum at 298 K both before and after heating (see Figure 5Ab and Bb).…”

Section: The Effect Of the Magnetic Fieldmentioning

confidence: 99%

See 1 more Smart Citation

The interaction of water molecules with purple membrane suspension using ²H double‐quantum filter, ¹H and ²H diffusion nuclear magnetic resonance

et al. 2004

View full text Add to dashboard Cite

Bacteriorhodopsin is a membrane protein of the purple membrane (PM) of Halobacterium salinarum, which is isolated as sheets of highly organized two-dimensional hexagonal microcrystals and for which water molecules play a crucial role that affects its function as a proton pump. In this paper we used single- and double-quantum (2)H NMR as well as (1)H and (2)H diffusion NMR to characterize the interaction of water molecules with the PM in D(2)O suspensions. We found that, under the influence of a strong magnetic field on a concentrated PM sample (0.61 mM), the PM sheets affect the entire water population and a residual quadrupolar splitting (upsilon(q) approximately 5.5 Hz, 298 K, at 11.7 T) is observed for the D(2)O molecules. We found that the residual quadrupolar coupling, the creation time in which a maximal DQF signal was obtained (tau(max)), and the relative intensity of the (2)H DQF spectrum of the water molecules in the PM samples (referred to herein as NMR order parameters) are very sensitive to temperature, dilution, and chemical modifications of the PM. In concentrated PM samples in D(2)O, these NMR parameters seem to reflect the relative organization of the PM. Interestingly, we have observed that some of these parameters are sensitive to the efficiency of the trimer packing, as concluded from the apo-membrane behavior. The data for dionized blue membrane, partially delipidated sample, and detergent-treated PM show that these D(2)O NMR order parameters, which are magnetic field dependent, are sensitive to the structural integrity of the PM. In addition, we revealed that heating the PM sample inside or outside the NMR magnet has, after cooling, a different effect on the NMR characteristics of the water molecules in the concentrated PM suspensions. The difference in the D(2)O NMR order parameters for the PM samples, which were heated and cooled in the presence and in the absence of a strong magnetic field, corroborates the conclusions that the above D(2)O order parameters are indirect reflections of both microscopic and macroscopic order of the PM samples. In addition, (1)H NMR diffusion measurements showed that at least three distinct water populations could be identified, based on their diffusion coefficients. These water populations seem to correlate with different water populations previously reported for the PM system.

show abstract

Thermal transitions in the purple membrane from Halobacterium halobium

Cited by 49 publications

References 24 publications

Thermal stability of lipid‐depleted purple membranes at neutral and low pH values

Thermal stability of lipid‐depleted purple membranes at neutral and low pH values

Irreversible thermal denaturation ofTorpedo californicaacetylcholinesterase

The interaction of water molecules with purple membrane suspension using ²H double‐quantum filter, ¹H and ²H diffusion nuclear magnetic resonance

Contact Info

Product

Resources

About

Thermal transitions in the purple membrane from Halobacterium halobium

Cited by 49 publications

References 24 publications

Thermal stability of lipid‐depleted purple membranes at neutral and low pH values

Thermal stability of lipid‐depleted purple membranes at neutral and low pH values

Irreversible thermal denaturation ofTorpedo californicaacetylcholinesterase

The interaction of water molecules with purple membrane suspension using 2H double‐quantum filter, 1H and 2H diffusion nuclear magnetic resonance

Contact Info

Product

Resources

About

The interaction of water molecules with purple membrane suspension using ²H double‐quantum filter, ¹H and ²H diffusion nuclear magnetic resonance