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W G Wu

Publications and source records attributed to W G Wu.

At least 19 recordsLinked to original sources
PubMed2001

Dynamic characterization of the water binding loop in the P-type cardiotoxin: implication for the role of the bound water molecule.

Recent studies of cobra P-type cardiotoxins (CTXs) have shown that the water-binding loop (loop II) plays a crucial role in toxin binding to biological membranes and in their cytotoxicity. To understand the role of bound water in the loop, the structure and dynamics of the major P-type CTX from Taiwan cobra, CTX A3, were determined by a comprehensive NMR analysis involving (1)H NOESY/ROESY, (13)C[1)H]NOE/T(1) relaxation, and (17)O triple-quantum filtered NMR. A single water molecule was found to be tightly hydrogen bonded to the NH of Met26 with a correlation time (5-7 ns) approaching the isotropic tumbling time (3.8-4.5 ns) of the CTX A3 molecule. Surprisingly, despite the relatively long residence time (ca. 5 ns to 100 micros), the bound water molecule of CTX A3 is located within a dynamic (order parameter S(2) approximately 0.7) and solvent accessible loop. Comparison among several P-type CTXs suggests that proline residues in the consensus sequence of MxAxPxVPV should play an important role in the formation of the water binding loop. It is proposed that the exchange rate of the bound water may play a role in regulating the lipid binding mode of amphiphilic CTX molecules near membrane surfaces.

Amino Acid Sequence↗
PubMed2000

Heparin reduces the alpha-helical content of cobra basic phospholipase A(2) and promotes its complex formation.

The interaction of phospholipase A(2) (PLA(2)) with glycosaminoglycans (GAGs) has recently attracted attention in view of its implication on inflammation and cell proliferation. By using Fourier Transformed Infrared (FTIR) spectroscopic measurements, we demonstrate here that binding of cobra basic phospholipase A(2) from Naja nigricollis (N-PLA(2)) to heparin may induce a significant conformational change observed in the amide I region of the enzyme's alpha-helical and beta-sheet structure. It is observed that notable conformational change of N-PLA(2) due to heparin binding occurs only when heparin's chain length is at least an octasaccharide as evidenced by circular dichroism and optical density measurements. This correlation may be an important factor in the aggregation of N-PLA(2) and N-PLA(2)-heparin complexes. Heparin induced change in conformation of PLA(2) is suggested to be a notable link in understanding the diversity in PLA(2) activity when rendered to the extracellular matrix of cell membranes that is full of GAG molecules.

Animals↗
PubMed2000

C-CAM1, a candidate tumor suppressor gene, is abnormally expressed in primary lung cancers.

Previous studies have shown that the expression of the cell-cell adhesion molecule (C-CAM1), located at chromosome 19, is down-regulated in several types of human cancers, including prostate and breast cancers. Two major isoforms of C-CAM1, the long or L-form C-CAM1 and the short or S-form C-CAM1, are derived from the C-CAM1 gene through alternative splicing. Tumor cells transfected with L-form C-CAM1, which contains a cytoplasmic domain, display significantly lower growth rates and less tumorigenicity in both in vitro and in vivo models compared with untransfected tumor cells, suggesting that L-form C-CAM1 may be a tumor suppressor. The transfection of the cytoplasmic domain of L-form C-CAM1 could also cause suppression of tumor growth, further supporting the role of L-form C-CAM1 in tumorigenesis. In contrast to reports of most of the tumor types tested, Ohwada et al. (Am. J. Respir. Cell Mol. Biol., 11: 214-220, 1994) reported that C-CAM1 was not down-regulated or even up-regulated in lung cancer. Because the cytoplasmic domain of L-form C-CAM1 is critical for the tumor suppressor function of C-CAM1, we hypothesized that switching of the isoform rather than down- regulation of C-CAM1 gene expression occurs during lung tumorigenesis. To test this hypothesis, we analyzed pairs of tumor tissue and corresponding normal-appearing lung tissue from 51 patients with non-small cell lung cancer (NSCLC) and 43 cell lines to determine expression profiles of L-form C-CAM1 and S-form C-CAM1 using reverse transcription-PCR. We found that L-form C-CAM1 was the predominant form (75%; 38 of 51) in normal-appearing lung tissue, whereas most (84%; 43 of 51) of the primary NSCLC tissue samples expressed predominantly S-form C-CAM1 (P < 0.0001). Similarly, 19 (79%) of the 24 NSCLC cell lines and 17 (85%) of the 20 small cell lung cancer cell lines expressed predominantly S-form C-CAM1. The frequent alteration of the C-CAM1 expression pattern suggests that C-CAM1 has an important role in lung tumorigenesis.

Adenosine Triphosphatases↗
PubMed1999

Heparin binding to cobra basic phospholipase A2 depends on heparin chain length and amino acid specificity.

Heparin is shown to bind specifically to the carboxy-terminal region of toxic type I phospholipase A2 from Naja nigricollis (N-PLA2) by competition assay using synthetic polypeptides and heparin affinity chromatography. The binding strength is seen to depend on heparin chain length and the presence of N-sulfate groups of heparin. It is observed that both electrostatic and non-electrostatic interactions are involved in the specific binding of heparin to the carboxy-terminus. When heparin's size is at least a decasaccharide, about two molecules of N-PLA2 bind to one molecule of heparin, as evidenced by the chemical estimate of protein to carbohydrate ratio in such N-PLA2/heparin complexes. Based on such a stoichiometric measurement and computer modeling of the N-PLA2/heparin complex, it is suggested that the binding sites of the two N-PLA2 molecules on one heparin molecule lie on the opposite sides of the heparin chain.

Amino Acid Sequence↗
PubMed1998

Cobra cardiotoxin and phospholipase A2 as GAG-binding toxins: on the path from structure to cardiotoxicity and inflammation.

Glycosaminoglycans (GAGs) represent the sulfated carbohydrate moieties of proteoglycans which occur abundantly in tissues of the cardiovascular system. Many proteins bind specifically to GAGs and perform an important role in inflammation, cell proliferation, and blood coagulation processes. Recently, in vitro GAG-binding studies of cardiotoxins (CTXs) and basic phospholipase A(2) (PLA(2)) from cobra venom established the toxins as two new families of GAG-binding proteins. In particular, discontinuous basic residues in beta-sheet CTXs may form a cationic cradle suitable for heparin binding, as in the case of fibronectin module III-13. The binding specificity of beta-sheet proteins to different GAGs can be further enhanced by involving other cationic clusters near the flexible loop of the molecule. Since the three-dimensional structures of many CTXs and PLA(2) are available, these two toxins may serve as models for the elucidation of the molecular recognition of GAG-binding proteins and also as polypeptide templates for further improvement of the binding specificity suitable for future biomedical application. Research along the line of GAG-guided toxicity of cobra venom components may help us to understand the functional role of GAGs and the action mechanism of cobra venom components in the cardiovascular system.

Journal Article↗
PubMed1997

Crystal structure of cardiotoxin V from Taiwan cobra venom: pH-dependent conformational change and a novel membrane-binding motif identified in the three-finger loops of P-type cardiotoxin.

The crystal structure of cardiotoxin V from Taiwan cobra venom (CTX A5) has been solved at pH 8.5 and refined to an R-factor of 20.7% for 7013 reflections [>2sigma(F)] between 8- and 2.19-A resolution. The refined model shows that CTX A5 exists as a dimer. The assembly consists of 974 non-hydrogen atoms from 124 residues and 73 water molecules. The global monomeric structure is similar to that determined by NMR at pH 3.7, characterized by a core formed by two beta-sheets connected with three-finger loops. However, local conformational differences are detected in two functionally important regions, loops I and II. A disparity between the NMR and X-ray structure of CTX A5 is detected near the tip of loop I and can be attributed to the difference in the protonation state of His4 at different pH, resulting in a reorientation of the His4 imidazole ring. A concerted motion of amino acid side chains located near His4 is detected and possibly contributes to the pH-dependent binding ability of CTX A5 to phospholipid model membranes. The second difference, detected at the tip of loop II, is due to the hydrophobic contact between CTX dimers in the crystal packing and the interaction of water molecules with amino acid residues in the loop II region of the CTX containing Pro31 (P-type CTX). This interaction forces loop II into a more rigid omega shape bridging the main chain at positions 27 and 34, contradictory to the flexible, tapering shape detected by NMR. Thus, a novel continuous hydrophobic column capable of binding to and possibly penetrating the membrane lipid bilayer is formed by the tips of the three-finger loops. In this respect, the X-ray crystal structure of CTX A5 may represent the CTX structure in the membrane-binding mode.

Amino Acid Sequence↗
PubMed1997

Membrane packing geometry of diphytanoylphosphatidylcholine is highly sensitive to hydration: phospholipid polymorphism induced by molecular rearrangement in the headgroup region.

Diphytanoylphosphatidylcholine (DPhPC) has often been used in the study of protein-lipid interaction and membrane channel activity, because of the general belief that it has high bilayer stability, low ion leakage, and fatty acyl packing comparable to that of phospholipid bilayers in the liquid-crystalline state. In this solid-state 31P and 2H NMR study, we find that the membrane packing geometry and headgroup orientation of DPhPC are highly sensitive to the temperature studied and its water content. The phosphocholine headgroup of DPhPC starts to change its orientation at a water content as high as approximately 16 water molecules per lipid, as evidenced by hydration-dependent 2H NMR study at room temperature. In addition, a temperature-induced structural transition in the headgroup orientation is detected in the temperature range of approximately 20-60 degrees C for lipids with approximately 8-11 water molecules per DPhPC. Dehydration of the lipid by one more water molecule leads to a nonlamellar, presumably cubic, phase formation. The lipid packing becomes a hexagonal phase at approximately 6 water molecules per lipid. A phase diagram of DPhPC in the temperature range of -40 degrees C to 80 degrees C is thus constructed on the basis of NMR results. The newly observed hydration-dependent DPhPC lipid polymorphism emphasizes the importance of molecular packing in the headgroup region in modulating membrane structure and protein-induced pore formation of the DPhPC bilayer.

Desiccation↗
PubMed1996

Conformational change and inactivation of membrane phospholipid-related activity of cardiotoxin V from Taiwan cobra venom at acidic pH.

The phospholipid binding activity of cardiotoxin V from Naja naja atra (CTX A5) was studied by use of Langmuir monolayers and found to exhibit pH-dependence in binding to phosphatidylcholine membrane with an apparent pKa around 6.0. Proton NMR investigation of the CTX A5 molecule in the presence of phosphatidylcholine micelles reveals a decrease in association of CTX A5 with membranes at low pH as a result of the protonation of His-4 near the membrane binding site of loop I region of CTX. The pH-dependent binding can be attributed mainly, but not solely, to the change in charge content of the CTX molecule upon His-4 protonation at the membrane/water interface. This is shown by analyzing the pH- and ionic strength dependence of binding of CTXs to phospholipid monolayers according to Gouy-Chapman theory. The protonation of the His-4 residue also results in a local conformational change in the loop I region since the chemical shifts of amide protons for the amino acid residues from Cys-3 to Thr-14 are all found to vary as a function of pH with an apparent pKa similar to that of His-4. Interestingly, the effect is relayed to other amino acid residues in the structural core of the protein such as those in C-terminal (Lys-60, Cys-61, and Asn-62) and triple-stranded antiparallel beta-sheet (Cys-22, Lys-24, Ala-25, Arg-38, and Ala-41) regions. An additional local conformational change in the molecule results around pH 5 as evidenced by circular dichroism spectroscopic studies, although this change does not affect the characteristic beta-sheet and three-finger loop structure of CTX molecule as revealed by two-dimensional NOESY 1H NMR study. The latter conformational change at acidic pH, however, completely inactivates CTX-induced aggregation/fusion activity of sphingomyelin vesicles. The results suggest that deciphering the functional sites of CTXs on the basis of structure and dynamics determined at low pH should be done with caution. Since 19 out of 44 CTX homologues with known amino acid sequence contain His-4, the effect of His-4 on the structure and function of CTX molecules is important and is discussed in terms of the diverse membrane targets of CTX subtypes. Also discussed is the pH-induced activation of snake venom proteins in the victim.

Animals↗
PubMed1996

The role of acidic amino acid residues in the structural stability of snake cardiotoxins.

We have recently shown that membrane-related activities of cardiotoxin V from Naja naja atra (CTX A5) are diminished at acidic pH although the overall beta-sheet structure of the molecule is maintained. In order to understand more about the mechanism of inactivation of CTX at acidic pH, we studied the effect of pH and denaturing reagents on the structural stability of CTX. We found, first, pH-induced structural transitions occurred in CTX A5 at two pH values as judged by the CD ellipticity around 195 nm: an increase in the beta-sheet content occurred around pH 4 and followed by a decrease, therein, around pH 2. The pKa of three acidic amino acid residues in CTX A5, i.e., Glu-17, Asp-42, and Asp-59, were determined to be 4.0, 3.2, and below 2.3, respectively, by NMR spectroscopy. The low pKa value of Asp-59 implies salt bridge formation between Lys-2 and Asp-59. Thus, electrostatic interaction may stabilize the three loop structure in addition to the hydrogen bonds between N- and C-termini of CTX molecule. Second, 2,2,2-trifluoroethanol (TFE) and guanidinium chloride (GdmHCI) were found to induce alpha-helical and random coil formation, respectively, in CTX A5 and eight other beta-sheet CTXs. Comparison of the relative potencies of TFE and GdmHCI to induce structural changes suggests that the amino acid residue located at position 17 plays a role in the structural stability. Specifically, CTXs containing negatively charged Glu-17 are least stable. It is suggested that Glu-17 may perturb the interaction between Lys-2 and Asp-59, and thus the overall stability of beta-sheet, in the presence of denaturing reagent. In conclusion, the perturbed structural stability of CTXs may partially explain the lower activity CTX exhibits at acidic pH. A structural model to account for the unfolding and refolding of CTX molecules without the breaking of disulfide bonds is also proposed.

Amino Acid Sequence↗
PubMed1996

Structure and dynamics of primary hydration shell of phosphatidylcholine bilayers at subzero temperatures.

Deuterium NMR relaxation and intensity measurements of the 2H-labeled H2O/dimyristoyl phosphatidylcholine bilayer were performed to understand the molecular origin of the freezing event of phospholipid headgroup and the structure and dynamics of unfrozen water molecules in the interbilayer space at subzero temperatures. The results suggest that about one to two water molecules associated with the phosphate group freeze during the freezing event of phospholipid headgroups, whereas about five to six waters near the trimethylammonium group behave as a water cluster and remain unfrozen at temperatures as low as -70 degrees C. In addition, temperature-dependent T1 and T2 relaxation times suggest that dynamic coupling occurs not only between the phosphate group and its bound water, but also between the methyl group and the adjacent water molecules. Based on these observations, the primary hydration shell of phosphatidylcholine headgroup at subzero temperatures is suggested to consist of two distinct regions: a clathrate-like water cluster, most likely a water pentamer, near the hydrophobic methyl group, and hydration water molecules associated with the phosphate group.

Deuterium↗
PubMed1995

A new method of superoxide free radical determination in ischemia and reperfusion injury of rat liver.

The present study was designed to utilize the chemiluminescence (CL) method for O2- analysis in ischemia and reperfusion of the whole rat liver. The results indicated that the O2- level was lower after 30 and 60 min ischemia. After 40 min reperfusion, the level of O2- increased obviously, reaching about 8.3-fold (30 min ischemia) and 9.0-fold (60 min ischemia) (P < 0.001, P < 0.01) respectively. The results also show that Chinese Traditional Medicines ligustrazine and salvia compound can scavenge O2- effectively.

Animals↗
PubMed1995

Solvent effect on phosphatidylcholine headgroup dynamics as revealed by the energetics and dynamics of two gel-state bilayer headgroup structures at subzero temperatures.

The packing and dynamics of lipid bilayers at the phosphocholine headgroup region within the temperature range of -40 to -110 degrees C have been investigated by solid-state nuclear magnetic resonance (NMR) measurements of selectively deuterium-labeled H2O/dimyristoylphosphatidylcholine (DMPC) bilayers. Two coexisting signals with 2H NMR quadrupolar, splittings of 36.1 and 9.3 (or smaller) kHz were detected from the -CD3 of choline methyl group. These two signals have been assigned to two coexisting gel-state headgroup structures with fast rotational motion of -CD3 and -N(CD3)3 group, respectively, with a threefold symmetry. The largest quadrupolar splitting of the NMR signal detected from the -CD2 of C alpha and C beta methylene segment was found to be 115.2 kHz, which is 10% lower than its static value of 128.2 kHz. Thus, there are extensive motions of the entire choline group of gel-state phosphatidylcholine bilayers even at a subzero temperature of -110 degrees C. These results strongly support the previous suggestion (E. J. Dufourc, C. Mayer, J. Stohrer, G. Althoff, and G. Kothe, 1992, Biophys. J. 61:42-57) that 31P chemical shift tensor elements of DMPC determined under similar conditions are not the rigid static values. The free energy difference between the two gel-state headgroup structures was determined to be 26.3 +/- 0.9 kJ/mol for fully hydrated bilayers. Furthermore, two structures with similar free energy difference were also detected for "frozen" phosphorylcholine chloride solution in a control experiment, leading to the conclusion that the two structures may be governed solely by the energetics of fully hydrated phosphocholine headgroup. The intermolecular interactions among lipids, however, stabilize the static headgroup structure as evidenced by the apparently lower free energy difference between the two structures for partially hydrated lipid bilayers. Evidence is also presented to suggest that one of the headgroup structures with trimethylammonium group rotation, which is not compatible with the static headgroup structure in crystals, is due to the dielectric relaxation of the slowly reorienting inter bilayer water molecules near the physical edge of membrane surface. Finally, a molecular model of the hydration-induced conformational changes at the torsion angle a5 of the O-C-CN+ bond is proposed to explain the two detected coexisting headgroup structures. These results emphasize the important role of the trimethylammonium group in monitoring the structure and dynamics of the lipid headgroup.

Deuterium↗
PubMed1995

Molecular order and hydration property of amine group in phosphatidylethanolamine and its N-methyl derivatives at subzero temperatures.

The molecular order and hydration properties of the amine group in phosphatidylethanolamine and its N-methyl derivatives were studied by 2H-NMR at subzero temperatures. Three coexisting signals with 2H-NMR quadrupolar splittings of 146, 106, and 28.8 KHz were detected from the fully hydrated phosphatidylethanolamine/D2O at the lowest studied temperature of -120 degrees C by using short recycle time in the applied NMR pulse sequence. These signals have been assigned to originate from frozen D2O in the interbilayer space and the deuterated amine group, i.e., -ND, with and without threefold symmetric motions. Comparative 2H-NMR studies of phosphatidylethanolamine/D2O with different degrees of methylation over a temperature range between -40 and -120 degrees C lead to the following conclusions. First, the bond angle of -D attached to the nitrogen atom of the amine group may be determined by the 2H-NMR quadrupolar splittings, i.e., 106 and 28.8 KHz, of the two coexisting signals of the deuterated amine group and found to be 112.9 for the gel-state phosphatidylethanolamine. Second, assuming the applicability of the empirical equation for the hydrogen bond distance of N+D--O with deuteron quadrupole coupling constants and using the intermolecular hydrogen bond distance of the amine group determined in single crystals of phosphatidylethanolamine bilayers, the largest measured quadrupolar splitting (delta nu Q) of N-D in this study, i.e., 106 KHz, is close to the static value. This interpretation is also consistent with the fact that the delta nu Q value determined remains constant in the temperature range between -70 and -120 degrees C. Third, the molecular order parameter of the amine group, as calculated from the ratio of the libration-averaged and static delta nu Q value for the lipid with different degrees of methylation, suggests that the perturbation of the headgroup interaction is most significant for the final methylation step. Finally, measurement of the spectral intensity of isotropic unfrozen D2O signals in D2O/phospholipid dispersions at temperatures below the homogeneous nucleation temperature of ice formation for D2O, i.e., below -34 degrees C, suggests that the first methylation step perturbs the neighboring water most significantly. Assuming that the molecular order of the amine group and the amount of unfrozen water detected under the present experimental condition can be taken as a measure of the hydrogen-bonding ability and the extent of perturbation caused by the methyl group, respectively, the gradual methylation of the amine group perturbs the interactions of the N-methylated headgroups in a nonlinear fashion. The results provide a molecular explanation for the phase behavior of phospholipids with different degrees of methylation.

Deuterium↗
PubMed1994

Expression of glutathione S-transferase-cardiotoxin fusion protein in Escherichia coli.

We report here the construction of cardiotoxin V gene, from cobra snake venom (Naja naja atra), by chemically synthesized oligonucleotides and its expression as a glutathione S-transferase-cardiotoxin fusion protein in the inclusion bodies of Escherichia coli. The expression of cardiotoxin fusion protein in protein with a yield of about 35 mg/liter culture was confirmed by highly specific anti-peptide antibodies generated against the unique amino acid residues located at the tip of loop II of cardiotoxin V. Since the fusion protein can be easily treated by CNBr to free the toxin moiety, as revealed by immunoblotting of the cleaved protein, the results provide an avenue for future structural and functional studies of cardiotoxin molecules.

Amino Acid Sequence↗
PubMed1993

Solution structure of cardiotoxin V from Naja naja atra.

Cardiotoxins are small proteins that are found in the venoms of snakes from the Elapidae family. These toxins are known to bind to and disrupt the organization, integrity, and function of the cell membrane. Most of the well-studied cardiotoxins cause depolarization of membrane potentials and/or lysis of red cells. In contrast, CTX V from Naja naja atra displays poor hemolytic activity but is proficient at inducing aggregation and fusion of sphingomyelin vesicles [Chien et al. (1991) J. Biol. Chem. 266, 3252-3259]. To determine whether the unique activity of this CTX is attributable to its tertiary structure, the solution structure of CTX V was determined by NMR methods. On the basis of these studies, this cardiotoxin has the same general topology as other members of the family, and thus its unusual properties do not arise from any gross structural differences that are detectable by solution NMR methods. Molecular dynamics calculations indicate that residues 36-50 show concerted fluctuations. On the basis of sequence similarity, we postulate that residues 30-34 are important in determining the specificity of cardiotoxins for fusion versus lysis of vesicles.

Amino Acid Sequence↗
PubMed1993

Aspartyl residue 10 is essential for ATPase activity of rat hsc70.

Three mutants of rat hsc70 were constructed, overexpressed in Escherichia coli, purified, and characterized. First, site-directed mutation was utilized to substitute Asn for Asp-10. The recombinant protein, hsc70(D10N), loses not only its peptide-stimulated ATPase activity but also its basal ATPase activity. The measured dissociation constants of ATP (0.3 microM) and S-peptide (5 microM) for hsc70(D10N), however, are virtually identical to those of hsc70. The intrinsic fluorescence spectra of hsc70(D10N) also remain largely unchanged. Therefore, the overall structure of the hsc70 protein is most likely intact after mutation. Second, the entire C-terminal peptide-binding domain was deleted and the resultant mutant contains only the N-terminal ATPase domain of hsc70. This recombinant protein, Nt-hsc70, is a peptide-independent ATPase. The ATPase activity at 37 degrees C of the Nt-hsc70, 270 pmol/h/micrograms of protein, is comparable to that of maximally peptide-activated hsc70. Third, the Asp-10 of Nt-hsc70 was replaced by Asn. Despite that this mutant, Nt-hsc70(D10N), is capable of binding ATP and it loses the capability to hydrolyze ATP. Taken together, these results indicate that aspartyl residue 10 of hsc70 is essential for ATP hydrolysis. Purified hsc70 and its mutants autophosphorylate in vitro at a substoichiometric level. On average, less than 1% of the hsc70 and Nt-hsc70 proteins are phosphorylated. Although the amount of phosphate incorporated into hsc70(D10N) and Nt-hsc70(D10) is reduced, a significant level of phosphorylation can still be achieved in these two site-directed mutants. Hence, autophosphorylation of hsc70 and its mutants is not correlated with their ability to hydrolyze ATP.

Adenosine Triphosphatases↗
PubMed1991

Freezing of phosphocholine headgroup in fully hydrated sphingomyelin bilayers and its effect on the dynamics of nonfreezable water at subzero temperatures.

Differential scanning calorimetry (DSC) and nuclear magnetic resonance (NMR) spectroscopy are applied to characterize the nonfreezable water molecules in fully hydrated D2O/sphingomyelin at temperatures below 0 degrees C. Upon cooling, DSC thermogram displays two thermal transitions peaked at -11 and -34 degrees C. The high-temperature exothermic transition corresponds to the freezing of the bulk D2O, and the low-temperature transition, which has not previously been reported, can be ascribed to the freezing of the phosphocholine headgroup in the lipid bilayer. The dynamics of nonfreezable water are also studied by 2H NMR T1 (spin-lattice relaxation time) and T2e (spin-spin relaxation time obtained by two pulse echo) measurements at 30.7 MHz and at temperatures down to -110 degrees C. The temperature dependence of the T1 relaxation time is characterized by a distinct minimum value of 2.1 +/- 0.1 ms at -30 degrees C. T2e is discontinuous at temperature around -70 degrees C, indicating another freezing-like event for the bound water at this temperature. Analysis of the relaxation data suggest that nonfreezable water undergoes both fast and slow motions at characteristic NMR time scales. The slow motions are affected when the lipid headgroup freezes.

Calorimetry, Differential Scanning↗