Biomedical subjects

A Dong

Publications and source records attributed to A Dong.

At least 37 records · Page 2Linked to original sources

PubMed1997

A lysine 73-->histidine variant of yeast iso-1-cytochrome c: evidence for a native-like intermediate in the unfolding pathway and implications for m value effects.

In this paper we report thermodynamic studies on a variant of yeast iso-1-cytochrome c in which a surface lysine residue at position 73 has been replaced with a histidine (H73). Guanidine hydrochloride denaturation studies monitored by circular dichroism spectroscopy indicated decreased thermodynamic stability (a lower delta G(o)(u)H20) and a smaller m value for the H73 protein as compared to the wild type (WT) protein. Further investigations to probe the causes for the thermodynamic stability differences between the two proteins involved guanidine hydrochloride and urea denaturations monitored by tryptophan fluorescence. The stability of heme ligation in the denatured state in the presence of either guanidine hydrochloride or urea was monitored by the spin-state transition of the heme iron induced by pH. None of these studies supported the hypothesis that the decreased m value was due to heme-His73 ligation in the denatured state. Guanidine hydrochloride denaturations monitored by the change in the extinction coefficient at 695 nm, which is sensitive to the presence of heme-Met80 ligation, revealed a native-like intermediate for the H73 protein, probably caused by displacement of the Met80 heme ligand by histidine 73 at guanidine hydrochloride concentrations much lower than required for full cooperative unfolding. Presence of the native-like intermediate is most likely the cause of the smaller m value and decreased thermodynamic stability for the CD-monitored H73 protein unfolding as compared to the unfolding of the WT protein. Guanidine hydrochloride denaturations in the presence of 200 mM imidazole provide further evidence in support of the proposed mechanism.

Circular Dichroism↗

PubMed1997

Drug delivery matrix containing native protein precipitates suspended in a poloxamer gel.

Sustained delivery systems can achieve more constant blood levels of protein therapeutics than those obtained with bolus doses, leading to improved drug efficacy and fewer adverse side effects. Several different polymeric delivery systems have been studied, including poloxamers, which are unique because they can be prepared in aqueous buffers that are compatible with proteins. Poloxamers are nontoxic block copolymers of poly(ethylene oxide) and poly(propylene oxide). Certain poloxamers exhibit reversible thermal gelation. Thus, a solution of protein and poloxamer prepared at low temperatures and injected extravascularly will form a gel as it warms to body temperature. Subsequently, the protein is released slowly from the gel. To date, however, poloxamer gel delivery systems have been limited to relatively low protein concentrations (i.e., < or = 0.4 mg/mL) that produce a completely soluble protein and an optically clear gel. Much higher concentrations of other protein drugs might be needed to obtain an efficacious sustained dose. In the current in vitro study we found that a poloxamer 407 (22% wt/wt) matrix could be prepared containing tens of milligrams/mililiter of the model proteins alpha-chymotrypsin and lactate dehydrogenase. Under these conditions the protein forms a homogeneous suspension. Warming through the poloxamer 407 transition temperature (ca. 18 degrees C) results in a gel that retains a homogeneous distribution of protein precipitates for several days at 37 degrees C. Infrared spectroscopy documented that the precipitated proteins in the suspension have native secondary structure. Furthermore, the fully active protein can be recovered completely when the gel is dissolved in excess buffer. Finally, at the higher protein concentrations used to form the suspensions in poloxamer 407, protein stability during incubation at 37 degrees C was greatly improved over that seen at lower protein concentrations.

Buffers↗

PubMed1996

Effect of secondary structure on the activity of enzymes suspended in organic solvents.

Despite the extensive use and study of enzymes suspended in organic solvents, whether activity differences between different preparations can be accounted for by differences in protein secondary structure is still unknown. To address this issue, in the current study two model enzymes, alpha-chymotrypsin and subtilisin Carlsberg, were lyophilized and suspended in both polar and nonpolar organic solvents. The secondary structures of the proteins in the initial aqueous solution, in the lyophilized powder, and in the subsequent suspensions in organic solvents were determined using infrared spectroscopy. Lyophilization perturbed the secondary structure of both enzymes. With alpha-chymotrypsin, lyophilization from buffer followed by suspension in ethanol, hexane, or pyridine did not alter the unfolded structure observed in the dried powder. In contrast, with subtilisin Carlsberg, suspension of the dried enzyme in ethanol led to further perturbation of structure, whereas in hexane, and more so in pyridine, there was some return toward native structure. Lyophilization of the aqueous protein solutions in the presence of either trehalose or sorbitol led to retention of more native-like structure of both enzymes in the dried solid. However, large structural perturbations arose when these samples were suspended in organic solvents. The only exception was the subtilisin-trehalose mixture, which regained some native structure in ethanol and hexane. The greatest changes were noted in samples suspended in pyridine, in which the infrared spectra indicated extensive intermolecular beta-sheet formation from protein aggregates. There was not any consistent correlation between activity in organic solvents and either the initial structure obtained in the dried powders or the final structure when suspended in organic solvents. Nor could differences in residual water contents in dried samples or the total water content in the organic solvent reaction system account for the activity differences.

Chymotrypsin↗

PubMed1996

Differences in conformational dynamics of ribonucleases A and S as observed by infrared spectroscopy and hydrogen-deuterium exchange.

Differences in conformational dynamics of bovine pancreatic RNase A and RNase S have been investigated using hydrogen-deuterium (H-D) exchange in conjunction with Fourier transform infrared spectroscopy. Deuteration-induced spectral changes in the amide I and II regions were monitored as a function of time. Second-derivative analysis revealed similar amide I spectral patterns for both proteins in H2O as well as fully deuterated in D2O. However, the rate of amide proton exchange of RNase S is much faster than that of RNase A at 25 degrees C as determined by changes in the intensity ratio of amide II/amide I bands and frequency red-shifts of amide I components. The frequency red-shifts of the amide I components ascribed to beta-sheet, alpha-helix, and beta-turns are continuous as a function of time, indicating that both proteins are too small to contain isolated secondary structural groups containing only exchanged or unexchanged amide protons in the partially deuterated intermediate states. Despite the dramatic difference in H-D exchange rate, the patterns of spectral changes in the conformation-sensitive amide I regions of RNase A and RNase S are very similar throughout the course of deuteration, indicating a similar pathway of amide proton exchange in both proteins.

Animals↗

PubMed1996

Physical factors affecting the storage stability of freeze-dried interleukin-1 receptor antagonist: glass transition and protein conformation.

The effects of glass transition of, and protein conformation in, the dried solid on the storage stability of freeze-dried recombinant human interleukin-1 receptor antagonist (rhIL-1ra) were examined. Glass transition is a temperature-dependent phenomenon. Amorphous materials become hard and brittle at temperatures below their characteristic glass transition temperatures (Tg) such that diffusion of molecules along the matrix is not sufficient to cause large-scale structural changes. To ascertain the importance of the glass transition in protein storage stability, we compared 10 different lyophilized rhIL-1ra formulations, with Tgs ranging from 20 to 56 degrees C, during several weeks of storage at temperatures above and below the samples' Tgs. Protein degradation, both deamidation and aggregation, was greatly accelerated at temperatures above Tg, but for some formulations also arose below Tg. Thus, storage of dried proteins below the Tg is necessary but not sufficient to ensure long-term stability. To examine the effects of protein structure in the dried solid, we prepared formulations with various sucrose concentrations, all of which had a Tg = 66 +/- 2.5 degrees C. With infrared spectroscopy, we determined that the protein lyophilized with </=1% sucrose was unfolded in the initial dried solid. In contrast, in those formulations with >/=5% sucrose, conformational change was inhibited during lyophilization. When stored at 50 degrees C, degradation of the freeze-dried protein varied inversely with sucrose concentration. These results indicate that structural changes arising during the lyophilization process led to damage during subsequent storage, even if the storage temperature was less than the Tg. Together the results of these studies document that to obtain optimum stability of dried rhIL-1ra it was necessary to inhibit conformational change during lyophilization and to store at temperatures below the Tg of the dried formulation.

Amides↗

PubMed1996

Infrared and circular dichroism spectroscopic characterization of structural differences between beta-lactoglobulin A and B.

Structural differences between two genetic variants of bovine beta-lactoglobulins (type A and B) in aqueous solutions were characterized using Fourier transform infrared and circular dichroism spectroscopies. To probe differences in structural dynamics, the effects hydrogen-deuterium exchange were also compared for the two proteins. The infrared spectra recorded in H2O solution for the two proteins were nearly identical in the conformationlly sensitive amide I region. The only exceptions were small differences at the band ascribed to a high-wavenumber beta-sheet component near 1693 cm-1 and the band assigned to turns at 1684 cm-1. In contrast, when the proteins were prepared in D2O solution, marked spectral differences were observed at all regions ascribed to beta-sheet and turn structures. These differences are consistent with the structural differences of the two variants at amino acid residues 64 and 118, which are located at a turn and a beta-sheet structure, respectively, as revealed by X-ray crystallographic studies [Monaco et al. (1987) J. Mol. Biol. 197, 695-706]. The circular dichroism spectra for the two proteins were essentially identical, both before and after hydrogen-deuterium exchange. Therefore, hydrogen-deuterium exchange did not alter the proteins' secondary structure. The enhancement of the amide I spectral difference upon hydrogen-deuterium exchange was ascribed to the differences in the structural mobility of the two proteins. Since the rate of exchange was greater for variant A, it was concluded that this variant has greater structural mobility than variant B. These findings indicate that the combination of infrared spectroscopy and hydrogen-deuterium exchange has great potential in characterization of even subtle structural differences in proteins induced by naturally occurring point mutations and/or site-directed mutagenesis.

Animals↗

PubMed1996

Counteracting effects of thiocyanate and sucrose on chymotrypsinogen secondary structure and aggregation during freezing, drying, and rehydration.

Studies of numerous proteins with infrared spectroscopy have documented that unfolding is a general response of unprotected proteins to freeze-drying. Some proteins that are unfolded in the dried solid aggregate during rehydration, whereas others refold. It has been proposed for the latter case that aggregation is avoided because refolding kinetically outcompetes intermolecular interactions. In contrast, with proteins that normally aggregate after rehydration, minimizing unfolding during freeze-drying with stabilizer has been shown to be needed to favor the recovery of native protein molecules after rehydration. The purpose of the current study was to examine first the opposite situation, in which a denaturant is used to foster additional unfolding in the protein population during freeze-drying. If the protein is not intrinsically resistant to aggregation under the study conditions (e.g., because of intermolecular charge repulsion) and the denaturant does not disrupt intermolecular interactions during rehydration, this treatment should favor aggregation upon rehydration. With infrared spectroscopy we found that at concentrations of the denaturant Na thiocyanate (NaSCN) that only slightly perturbed chymotrypsinogen secondary structure in solution before freeze-drying, there was a large increase in protein unfolding in the dried solid and in protein aggregation measured after rehydration. Bands assigned to intermolecular beta sheet were present in the spectra of samples dried with NaSCN, indicating that aggregation could also arise in the dried solid. By examining the protein structure in the frozen state, we determined that in the absence of NaSCN the protein remains native. NaSCN caused structural perturbations during freezing, without the formation of intermolecular beta sheet, that were intermediate to structural changes noted after freeze-drying. In contrast, samples treated in the presence of NaSCN and sucrose had native-like spectra in the frozen and dried states, and much reduced aggregation after rehydration. These results indicate that during freezing and drying the sugar can counteract and mostly reverse the structural perturbations induced by NaSCN before and during these treatments.

Animals↗

PubMed1996

Formation of an active dimer during storage of interleukin-1 receptor antagonist in aqueous solution.

The degradation products of recombinant human interleukin-1 receptor antagonist (rhIL-1ra) formed during storage at 30 degrees C in aqueous solution were characterized. Cationic exchange chromatography of the stored sample showed two major, new peaks eluting before (P1) and after (L2) the native protein, which were interconvertible. Size-exclusion chromatography and electrophoresis documented that both the P1 and L2 fractions were irreversible dimers, formed by noncovalent interactions. A competition assay with interleukin-1 indicated that on a per monomer basis the P1 and L2 dimers retained about two-thirds of the activity of the native monomer. Infrared and far-UV circular dichroism spectroscopies showed that only minor alterations in secondary structure arose upon the formation of the P1 dimer. However, alteration in the near-UV circular dichroism spectrum suggested the presence of disulfide bonds in the P1 dimer, which are absent in the native protein. Mass spectroscopy and tryptic mapping, before and after carboxymethylation, demonstrated that the P1 dimer contained an intramolecular disulfide bond between Cys-66 and Cys-69. Although conversion of native protein to the P1 dimer was irreversible in buffer alone, the native monomer could be regained by denaturing the P1 dimer with guanidine hydrochloride and renaturing it by dialysis, suggesting that the intramolecular disulfide bond does not interfere with refolding. Analysis of the time course of P1 formation during storage at 30 degrees C indicated that the process followed first-order, and not second-order, kinetics, suggesting that the rate-limiting step was not dimerization. It is proposed that a conformational change in the monomer is the rate-limiting step in the formation of the P1 dimer degradation product. Sucrose stabilized the native monomer against this process. This result can be explained by the general stabilization mechanism for this additive, which is due to its preferential exclusion from the protein surface.

Anilino Naphthalenesulfonates↗

PubMed1996

Quantitation of the area of overlap between second-derivative amide I infrared spectra to determine the structural similarity of a protein in different states.

Maintaining a native-like structure of protein pharmaceuticals during lyophilization is an important aspect of formulation. Infrared spectroscopy can be used to evaluate the effectiveness of formulations in protecting the secondary structural integrity of proteins in the dried solid. This necessitates making quantitative comparisons of the overall similarity of infrared spectra in the conformationally sensitive amide I region. We initially used the correlation coefficient r, as defined by Prestrelski et al. (Biophys. J. 1993, 65, 661-671), for this quantitation. Occasionally, we noticed that the r value did not agree with a visual assessment of the spectral similarity. In some cases this was due to an offset in baselines, which led artifactually to an unreasonably low r value. Conversely, if the spectra were baseline corrected and there existed a large similarity between peak positions, but differences in relative peak heights, the r value would be unreasonably high. Our approach to avoiding these problems is to use area-normalized second-derivative spectra. We have found that quantitating the area of overlap between area-normalized spectra provides a reliable, objective method to compare overall spectral similarity. In the current report, we demonstrate this method with selected protein spectra, which were taken from experiments where unfolding was induced by lyophilization or guanidine hydrochloride, and artificial data sets. With this analysis, we document how problems associated with calculation of the correlation coefficient, r, are avoided.

Amides↗

PubMed1996

Antineoplastic agents; I. Three spirostanol glycosides from rhizomes of Dioscorea collettii var. hypoglauca.

By activity-guided fractionation, three known steroidal saponins, prosapogenin A of dioscin, dioscin and gracillin, were isolated from the total saponin fraction of Dioscorea coiletti var. hypoglauca as active compounds causing morphological abnormality of Pyricularia oryzae mycelia. The compounds also exhibited cytotoxic activity against the cancer cell line K562 in vitro. The structures of the compounds were elucidated on the basis of chemical evidence and IR, FAB-MS, 1H-NMR, 13C-NMR, and two-dimensional NMR (2D-NMR) analysis.

Antineoplastic Agents↗

PubMed1995

Redox-dependent changes in beta-sheet and loop structures of Cu,Zn superoxide dismutase in solution observed by infrared spectroscopy.

Redox-dependent conformational changes of bovine Cu,Zn superoxide dismutase in 20 mM phosphate buffer (pH 7.4) were studied at 20 degrees C using Fourier transform infrared spectroscopy. Amide I spectra provide evidence that conformational changes in the protein accompany a change in the oxidation state of copper at the active site. Quantitative analysis of these spectra indicates that both reduced (CuI,ZnII) and oxidized (CuII,ZnII) enzymes are composed of about 35% antiparallel beta-sheet, 45% unordered/loop, and 20% beta-turn structures. Significant redox-dependent changes occur in regions ascribed to beta-sheet and unordered/loop structures that are consistent with an active channel structure wherein the copper ion bonds to imidazolate side chains of His 44, 46, and 118 within the beta-sheet structure and also to the imidazolate side chain of His 61 associated with unordered/loop structure. This study provides the first experimental evidence that an unordered structure can exhibit bands in more than one region, one near 1658 cm-1 and another near 1648 cm-1 in both H2O and D2O solutions. The detected changes in protein conformation are expected to be critical to the catalytic function of this enzyme.

Animals↗

PubMed1995

Biochemical characterization of basilase, a fibrinolytic enzyme from Crotalus basiliscus basiliscus.

Snake venoms, especially from the Crotalidae family, contain a variety of enzymes that prevent blood coagulation by virtue of their fibrinolytic enzymes. Nineteen snake venoms were screened for fibrinolytic activity and the highest activity was found in the venom of Crotalus basiliscus basiliscus venom. The active principle, basilase, was isolated, purified, and found to have fibrinolytic and fibrinogenolytic activity. It had a molecular weight of 22,000 and 1 mol of zinc per mole of protein associated with it. The proteolytic activity of the enzyme against dimethyl casein was inhibited by ethylenediaminetetraacetic acid and alpha 2-macroglobulin. It did not inactivate alpha 2-macroglobulin. Basilase did not have any of the following activities: thrombin-like, factor X-like, protein C activating, or urokinase-like. It caused neither hemorrhage nor platelet aggregation. In spite of its proteolytic activity, basilase did not hydrolyze the membranes of platelets. Basilase had 24% alpha-helix, 31% beta-sheet, 25% turns, and 20% unordered structure, as determined by Fourier Transform Infrared spectroscopy. Basilase is an enzyme that hydrolyzes fibrin directly without activation of plasminogen.

Amino Acid Sequence↗

PubMed1995

The effects of hydrophilic to hydrophobic surface mutations on the denatured state of iso-1-cytochrome c: investigation of aliphatic residues.

A series of hydrophilic to hydrophobic surface mutations were prepared at the highly solvent-exposed lysine 73 of iso-1-cytochrome c to assess the ability of such mutants to affect the energetics of the denatured state. In this report, the aliphatic hydrophobics (leucine, isoleucine, valine, alanine, glycine) were studied. The thermodynamic stability of each of these mutants was determined by guanidine hydrochloride denaturation. Both the free energy of unfolding in the absence of denaturant, delta GouH2O, and the slope, m, of a plot of the free energy of unfolding, delta Gou, versus [guanidine hydrochloride] show significant negative correlations with the 1-octanol to water transfer free energy, delta Gtr, of the amino acid side chain at position 73. A negative correlation with hydrophobicity is consistent with these mutants leading to more extensive hydrophobic clustering in the denatured state, consistent with the predictions of heteropolymer theory for compact denatured states; an effect operating on the native state energetics should produce a positive correlation of delta GouH2O with hydrophobicity. Infrared amide I spectroscopy indicated native state structural perturbations for the glycine 73 and isoleucine 73 mutants. A moderate correlation of delta GouH2O was also found with alpha-helix propensity, suggesting that both hydrophobic effects acting on the denatured state and alpha-helix propensity are affecting the delta GouH2O values for these mutants.

Amino Acids↗

PubMed1995

Infrared analysis of ligand- and oxidation-induced conformational changes in hemoglobins and myoglobins.

Effects of the binding of O2 and CO to heme iron (II) of deoxy forms and of the oxidation of deoxy forms to aquoiron (III) complexes on the infrared spectra of hemoglobins and myoglobins have been examined. Spectra were measured for aqueous solutions 3-4 mM in heme of human, bovine, and equine hemoglobins and sperm whale, bovine, and equine myoglobins in 10 mM sodium phosphate buffer, pH 7.4, at 20 degrees C. All ligand binding and oxidation reactions resulted in similar spectral shifts in the region 1665 to 1670 cm-1, a portion of the amide I region assignable to beta-turn structure. There were no other significant changes in the amide I region, a finding consistent with no other alterations in secondary structure. The major bands near 1655 cm-1 associated with alpha-helices were consistently at 2 cm-1 lower wavenumber for myoglobins than for hemoglobins. The changes in solution infrared spectra observed in this study may result at least in part from conformational changes at the FG corner associated with movements of F and E helices that have been noted previously in crystal structures.

Animals↗

PubMed1995

Effects of dimethyl sulfoxide, glycerol, and ethylene glycol on secondary structures of cytochrome c and lysozyme as observed by infrared spectroscopy.

Effects of 10-30% (v/v) of dimethyl sulfoxide, glycerol, and ethylene glycol on the H-O-H bending vibration of water and the amide I bands of horse heart cytochrome c and chicken egg white lysozyme in 25 mM sodium phosphate buffer (pH 7.4) were examined at 20 degrees C by Fourier transform infrared spectroscopy. The H-O-H bending mode of water was strongly affected by these cryoprotectant solvents. Increasing the concentration of cryosolvents from 0 to 30% shifts the water bending band maximum from 1645 to about 1650 cm-1. Second-derivative analysis reveals significant changes in conformation-sensitive amide I regions of lysozyme ascribed to alpha-helix (1657 cm-1), turn (1674 cm-1), and unordered (1646 cm-1) structures; each cryosolvent increases the intensity of the 1657 cm-1 band at the expense of bands at 1674 and 1646 cm-1. No changes in spectra deemed significant were observed for cytochrome c under the same conditions. There is no spectral evidence of structural randomization of proteins due to the presence of these cryosolvents. Cryosolvent-induced changes in secondary structure of proteins may result from changes in water structure which, in turn, perturb the structure of the protein and/or from direct interactions between cryosolvent and protein.

Animals↗

PubMed1995

Infrared spectroscopic studies of lyophilization- and temperature-induced protein aggregation.

Recent studies have clearly demonstrated that Fourier transform IR spectroscopy can be a powerful tool for the study of protein stabilization during freeze-drying and for optimizing approaches to prevent lyophilization-induced protein aggregation. The purpose of the current review is to provide an overview of these topics, as well as an introduction to the study of protein secondary structure with IR spectroscopy. We will start with a general summary of the theories and practices for processing and interpreting protein IR spectra. We will then review the current literature on the use of IR spectroscopy to study protein structure and the effects of stabilizers during lyophilization. Next we will concentrate specifically on protein aggregation. The bulk of the research and the key assignments of spectral features in protein aggregates come from studies of the effects of high and low temperature on proteins. Therefore, we will first consider this topic. Finally, we will summarize the recent theoretical and applied work on lyophilization-induced aggregation.

Chymotrypsin↗

PubMed1994

Determination of the amounts and oxidation states of hemoglobins M Boston and M Saskatoon in single erythrocytes by infrared microspectroscopy.

The reduced abnormal subunits of two M-type hemoglobins, Boston (His alpha 58-->Tyr) and Saskatoon (His beta 63-->Tyr), have been determined in the presence of normal human hemoglobin A by measurement of C-O stretch bands in infrared spectra of carbon monoxide complexes. Use of an infrared microscope coupled to a Fourier transform infrared spectrometer of high sensitivity permitted measurements to be made on as small a hemoglobin mixture as is contained in a single erythrocyte. The abnormal subunits of both Hbs M exhibit bands near 1970 cm-1 compared with bands near 1951 cm-1 for the normal subunits. The increase in 1970 cm-1 band intensity upon erythrocyte reduction with dithionite provided a measure of the extent of abnormal subunit oxidation; in cell suspensions about 60% of the abnormal subunits of Hb M Boston and 80% for Hb M Saskatoon remained reduced. The amount of Hb present as abnormal Hb averaged about 25% for Hb M Boston cells and about 50% for Hb M Saskatoon cells. However, the ratio of Hb M to Hb A in individual cells varied markedly, with the ratio expected to decrease as the cell ages. These results demonstrate the unique utility of infrared microspectroscopy for the study of differences in abnormal Hb status among individual erythrocytes.

Carboxyhemoglobin↗

PubMed1994

Characterization of sites occupied by the anesthetic nitrous oxide within proteins by infrared spectroscopy.

We report here a comprehensive infrared spectroscopic study of the interactions between the anesthetic nitrous oxide (N2O) and six proteins: lysozyme, cytochrome c, myoglobin, hemoglobin, serum albumin, and cytochrome c oxidase. Sites occupied by N2O molecules within these proteins were characterized. Three types of hydrophobic sites were found within the proteins. One with nu 3 near 2225 cm-1 is likely to be near peptide bond carbonyls; one with nu 3 near 2219 cm-1 may be near a benzene-like structure such as the side chains of phenylalanine and tyrosine; and the other with nu 3 near 2215 cm-1 is likely to be in a nonpolar alkane-like environment provided by the side chains of Leu, Ile, and Val residues. The amount of N2O molecules bound to myoglobin increases as the pH decreases from 9.2 to 5.2. N2O-protein interactions produced no detectable changes in the ligand-binding pockets of myoglobin, hemoglobin, and cytochrome c oxidase. N2O-induced secondary structure changes were detected only in the fully reduced cytochrome c oxidase, not in the fully oxidized oxidase and the other five proteins. N2O-induced conformational changes in the alpha beta-interface of hemoglobin and the h2 and h3 alpha-helices of human serum albumin were detected by monitoring the S-H stretch vibrations of cysteine residues. These findings provide direct evidence that anesthetic N2O interacts with proteins and occupies sites in the interior of the proteins.

Adult↗