Membrane stabilization during freezing: The role of two natural cryoprotectants, trehalose and proline

Rudolph, Alan S.; Crowe, John H.

doi:10.1016/0011-2240(85)90184-1

Cited by 385 publications

(179 citation statements)

References 24 publications

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“…In particular, proline and trehalose appear to bind to head groups of membrane phospholipids, in effect replacing water molecules. Thus, they can stabilize membranes during cell shrinkage (Rudolph and Crowe, 1985;Storey and Storey, 1996).…”

Section: Freezingmentioning

confidence: 99%

Organic osmolytes as compatible, metabolic and counteracting cytoprotectants in high osmolarity and other stresses

Yancey

2005

Journal of Experimental Biology

1,553

1,272

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SUMMARY Organic osmolytes are small solutes used by cells of numerous water-stressed organisms and tissues to maintain cell volume. Similar compounds are accumulated by some organisms in anhydrobiotic, thermal and possibly pressure stresses. These solutes are amino acids and derivatives,polyols and sugars, methylamines, methylsulfonium compounds and urea. Except for urea, they are often called `compatible solutes', a term indicating lack of perturbing effects on cellular macromolecules and implying interchangeability. However, these features may not always exist, for three reasons. First, some of these solutes may have unique protective metabolic roles, such as acting as antioxidants (e.g. polyols, taurine, hypotaurine),providing redox balance (e.g. glycerol) and detoxifying sulfide (hypotaurine in animals at hydrothermal vents and seeps). Second, some of these solutes stabilize macromolecules and counteract perturbants in non-interchangeable ways. Methylamines [e.g. trimethylamine N-oxide (TMAO)] can enhance protein folding and ligand binding and counteract perturbations by urea (e.g. in elasmobranchs and mammalian kidney), inorganic ions, and hydrostatic pressure in deep-sea animals. Trehalose and proline in overwintering insects stabilize membranes at subzero temperatures. Trehalose in insects and yeast,and anionic polyols in microorganisms around hydrothermal vents, can protect proteins from denaturation by high temperatures. Third, stabilizing solutes appear to be used in nature only to counteract perturbants of macromolecules,perhaps because stabilization is detrimental in the absence of perturbation. Some of these solutes have applications in biotechnology, agriculture and medicine, including in vitro rescue of the misfolded protein of cystic fibrosis. However, caution is warranted if high levels cause overstabilization of proteins.

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Section: Freezingmentioning

confidence: 99%

Organic osmolytes as compatible, metabolic and counteracting cytoprotectants in high osmolarity and other stresses

Yancey

2005

Journal of Experimental Biology

1,553

1,272

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“…In another study, trehalose was also found to be more efficient than glycerol and dimethylsulfoxide in the stabilization of sarcoplasmic reticulum during freeze-thaw. 8 Here we provide a method for the long-term storage of frozen mouse liver mitochondria in trehalose-containing buffer and show that trehalose-frozen mitochondria retain a number of their important biological functions, most notably the preservation of MOM integrity.…”

mentioning

confidence: 98%

Mitochondria frozen with trehalose retain a number of biological functions and preserve outer membrane integrity

et al. 2006

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In apoptosis, Bcl-2-family proteins regulate the barrier function of the mitochondrial outer membrane (MOM), controlling the release of proapoptotic proteins from the intermembrane space into the cytoplasm. This process can be studied in vitro with freshly isolated mouse liver mitochondria. Unfortunately, mitochondria frozen/thawed in standard sucrose-mannitol buffers become leaky and useless for apoptosis research. However, here we show that mitochondria frozen in buffer containing the sugar, trehalose, maintained MOM integrity and responsiveness to Bcl-2-family proteins, much like fresh mitochondria. Trehalose also preserved ultrastructure, as well as biological functions such as ATP synthesis, calcium-induced swelling, transmembrane potential, and the import and processing of protein precursors. However, bioenergetic function was somewhat reduced. Thus, trehalose-frozen mitochondria retained most of the biological features of mitochondria including MOM integrity. Although not ideal for studies involving bioenergetics, this method will facilitate research on apoptosis and other mitochondrial functions that rely on an intact MOM. Since the 1970s, there have been a number of attempts to freeze isolated mitochondria for storage, with various degrees of success. In most of these studies, the functionality of frozen-thawed mitochondria was assessed with regard to bioenergetic parameters. In a recent and relatively successful attempt, frozen mitochondria were reported to have roughly 50% of normal respiratory function, and the mitochondria were well coupled.

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“…Thus, studies on intact cells have documented improved survival following exposure to heat, cold, drought, or chemical stressors (6,10,11). Other works have analyzed stabilization on the basis of phenomenological properties of model membranes, for example, the leakage or intermixing of probes in liposomes (12,13). Finally, stability has been discussed with respect to rigorous physical parameters such as the structure or mechanical properties of lipid bilayers (14,15).…”

mentioning

confidence: 99%

Reconciliation of opposing views on membrane–sugar interactions

Andersen

Wang

Arleth

et al. 2011

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

133

177

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It is well established that small sugars exert different types of stabilization of biomembranes both in vivo and in vitro. However, the essential question of whether sugars are bound to or expelled from membrane surfaces, i.e., the sign and size of the free energy of the interaction, remains unresolved, and this prevents a molecular understanding of the stabilizing mechanism. We have used smallangle neutron scattering and thermodynamic measurements to show that sugars may be either bound or expelled depending on the concentration of sugar. At low concentration, small sugars bind quite strongly to a lipid bilayer, and the accumulation of sugar at the interface makes the membrane thinner and laterally expanded. Above ∼0.2 M the sugars gradually become expelled from the membrane surface, and this repulsive mode of interaction counteracts membrane thinning. The dual nature of sugar-membrane interactions offers a reconciliation of conflicting views in earlier reports on sugar-induced modulations of membrane properties.membrane interface | membrane structure | preferential binding | preferential exclusion | interaction free energy S mall sugars such as the disaccharides sucrose and trehalose are among the so-called osmolytes (1) or compensatory solutes (2), which are accumulated in response to environmental stress in virtually all taxa. Their function is to act as inert regulators of the osmotic pressure, but they also optimize the physical properties of the cytosol (3) and stabilize biomolecular conformations against cold, drought, and heat (4-7). The same small carbohydrates have also proven useful in vitro as protectants or excipients for biopreservation (8). Many reports have shown that membranous structures are particularly stabilized by small sugars (4, 6, 9), but the definition of stabilization covers a wide range of biological and physical parameters. Thus, studies on intact cells have documented improved survival following exposure to heat, cold, drought, or chemical stressors (6,10,11). Other works have analyzed stabilization on the basis of phenomenological properties of model membranes, for example, the leakage or intermixing of probes in liposomes (12, 13). Finally, stability has been discussed with respect to rigorous physical parameters such as the structure or mechanical properties of lipid bilayers (14, 15). The current work addresses membrane dimensions and the thermodynamics of interaction with the purpose of elucidating fundamental aspects of membrane-sugar interrelationships. The different observations of sugar stabilization have sparked a large number of studies on sugars and model membranes (usually phospholipid bilayers) over the past 30 y. Investigations of fully hydrated membranes show an interesting tendency to fall into two groups with mutually conflicting conclusions. Thus, many investigations have suggested direct (favorable) interaction of sugars and the phospholipid interface (16-23), and it is obvious that such interactions could be the origin of sugar effects, for example, through int...

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Membrane stabilization during freezing: The role of two natural cryoprotectants, trehalose and proline

Cited by 385 publications

References 24 publications

Organic osmolytes as compatible, metabolic and counteracting cytoprotectants in high osmolarity and other stresses

Organic osmolytes as compatible, metabolic and counteracting cytoprotectants in high osmolarity and other stresses

Mitochondria frozen with trehalose retain a number of biological functions and preserve outer membrane integrity

Reconciliation of opposing views on membrane–sugar interactions

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