Interaction of antimycobacterial and anti-pneumocystis drugs with phospholipid membranes

Liposomes can be used as carriers of drugs in the treatment of viral, bacterial and protozoal infections. The potential for liposome-mediated therapy of Mycobacterium avium-intracellulare complex infections, one of the most common opportunistic infections in AIDS, is currently under study. Here, we have investigated the effect of the lipid-soluble antimycobacterial drugs ansamycin, clofazimine and CGP7040 on the thermotropic behavior of liposomes composed of dipalmitoylphosphatidylcholine (DPPC) or dipalmitoylphosphatidylglycerol (DPPG) using differential scanning calorimetry (DSC). In the presence of ansamycin (rifabutine), the peak gel-liquid crystalline phase transition temperature (Tm) of DPPG was reduced, as was the sub-transition temperature (Ts), whereas the Tm of DPPC was reduced only slightly. The temperature of the pre-transition (Tp) of DPPC was lowered, while the pre-transition of DPPG was abolished. Ansamycin also caused the broadening of the transition endotherm of both lipids. Equilibration of the drug/lipid complex for 1 or 5 days produced different thermotropic behavior. In the presence of clofazimine, the cooperativity of the phase transition of DPPG decreased. Above 10 mol% clofazimine formed two complexes with DPPG, as indicated by two distinguishable peaks in DSC thermograms. The Tm of both peaks were lowered as the mole fraction increased. Clofazimine had minimal interaction with DPPC. In contrast, CGP7040 interacted more effectively with DPPC than with DPPG, causing a reduction of the size of the cooperative unit of DPPC even at 2 mol%. The main transition of DPPC split into 3 peaks at 5 mol% drug. The pre-transition was abolished at all drug concentrations and the sub-transition disappeared at 10 mol% CGP7040. These studies suggest that maximal encapsulation of clofazimine in liposomes would require a highly negatively charged membrane, while that of CGP7040 would necessitate a zwitterionic membrane. We have also investigated the interaction of the water-soluble antibiotic pentamidine, which has been used against Pneumocystis carinii, the most lethal of AIDS-related opportunistic pathogens. Aerosol administration of this drug leads to long-term sequestration of the drug in the lungs. The DPPG/pentamidine complex exhibited a pre-transition at 3.5 degrees C, an endothermic peak at 42 degrees C, and an exothermic peak at 44.5 degrees C, followed by another endothermic peak at 55 degrees C. The exotherm depended on the history of the sample, requiring pre-incubation for several minutes below the 42 degrees C transition. These observations suggest that upon melting of the DPPG chains at 42 degrees C, the DPPG crystallizes as a DPPG/pentamidine complex that melts at 55 degrees C.     

Susceptibility of cartilage collagens type II, IX, X, and XI to human synovial collagenase and neutrophil elastase

The action of purified rheumatoid synovial collagenase and human neutrophil elastase on the cartilage collagen types II, IX, X and XI was examined. At 25 degrees C, collagenase attacked type II and type X (45-kDa pepsin-solubilized) collagens to produce specific products reflecting one and at least two cleavages respectively. At 35 degrees C, collagenase completely degraded the type II collagen molecule to small peptides whereas a large fragment of the type X molecule was resistant to further degradation. In contrast, collagen type IX (native, intact and pepsin-solubilized type M) and collagen type XI were resistant to collagenase attack at both 25 degrees C and 35 degrees C even in the presence of excess enzyme. Mixtures of type II collagen with equimolar amounts of either type IX or XI did not affect the rate at which the former was degraded by collagenase at 25 degrees C. Purified neutrophil elastase, shown to be functionally active against soluble type III collagen, had no effect on collagen type II at 25 degrees C or 35 degrees C. At 25 degrees C collagen types IX (pepsin-solubilized type M) and XI were also resistant to elastase, but at 35 degrees C both were susceptible to degradation with type IX being reduced to very small peptides. Collagen type X (45-kDa pepsin-solubilized) was susceptible to elastase attack at 25 degrees C and 35 degrees C as judged by the production of specific products that corresponded closely with those produced by collagenase. Although synovial collagenase failed to degrade collagen types IX and XI, all the cartilage collagen species examined were degraded at 35 degrees C by conditioned culture medium from IL1-activated human articular chondrocytes. Thus chondrocytes have the potential to catabolise each cartilage collagen species, but the specificity and number of the chondrocyte-derived collagenase(s) has yet to be resolved.     

Structural diversity and domain composition of a unique collagenous fragment (intima collagen) obtained from human placenta.

Intima collagen was obtained from pepsin digests of human placenta in two forms, which differ to some extent in the size of their constituent polypeptide chains (Mr 50 000-70 000). These chains are connected by disulphide bonds to large aggregates. The aggregates are arranged in a triple-helical conformation with a remarkably high thermal stability (Tm 41-62 degrees C) and are resistant to further proteolytic digestion. Reduction of as little as 5% of the disulphide bonds produces mainly monomeric triple helices (Mr about 160 000) with Tm 32 degrees C. Partially reduced material can be separated into triple-helical and non-collagenous domains by proteolysis. Pepsin releases a collagenous component with chains of Mr 38 000. Bacterial collagenase liberates two non-collagenous segments (Mr 15 000-30 000) rich in cystine. Treatment with collagenase before reduction separates intima collagen into a large fragment composed of collagenous (Tm 41 degrees C) and non-collagenous structures and a single non-collagenous segment. The data support the electron-microscopical model of intima collagen [Furthmayr, Wiedemann, Timpl, Odermatt & Engel (1983) Biochem. J. 211, 303-311], indicating that the basic unit of the fragment consists of a continuous triple helix joining two globular domains.     

Cleavage of bovine skin type III collagen by proteolytic enzymes. Relative resistance of the fibrillar form

We have studied the susceptibility of fibrils formed from fetal bovine skin type III collagen to proteolytic enzymes known to cleave within the helical portion of the molecule (vertebrate and microbial collagenase, polymorphonuclear elastase, trypsin, thermolysin) and to two general proteases of broad specificity (plasmin, Pronase). Fibrils reconstituted from neutral salt solutions, at 35 degrees C, were highly resistant to nonspecific proteolysis by general proteases such as polymorphonuclear elastase, trypsin, and thermolysin but were rapidly dissolved by bacterial and vertebrate collagenases at rates of 12-45 mol X mol-1 X h-1. In solution, type III collagen was readily cleaved by each of the proteases (with the exception of plasmin), as well as by the true collagenases, although at different rates. Turnover numbers determined by viscometry at 35 degrees C were: human collagenase, approximately equal to 1500 h-1; microbial (clostridial) collagenase, approximately equal to 100 h-1; and general proteases, 23-52 h-1. In addition it was shown that pronase cleaves type III collagen in solution at 22 degrees C by attacking the same Arg-Gly bond in the alpha 1(III) chain as trypsin. However, like other proteases, Pronase was rather ineffective against fibrillar forms of type III collagen. It was also shown that transition of type III collagen as well as type I collagen to the fibrillar form resulted in a significant gain of triple helical thermostability as evidenced by a 6.8 degrees C increase in denaturation temperature (Tm = 40.2 degrees C in solution; Tm = 47.0 degrees C in fibrils).     

Collagen from diamondback squid (Thysanoteuthis rhombus) outer skin

Collagens (acid-solubilized and pepsin-solubilized collagens) were prepared from diamondback squid outer skin and partially characterized. The yields of acid-solubilized and pepsin-solubilized collagens were about 1.3 and 35.6%, respectively, on a dry weight basis. Pepsin-solubilized collagen was heterotrimer with a chain composition of ala2a3. The patterns of peptide fragments were different from that of porcine skin collagen. Denaturation temperature was 27.5 degrees C, about 10 degrees C lower than that of porcine collagen. The amino acid composition of pepsin-solubilized collagen from diamondback squid outer skin was similar to that from cuttlefish outer skin. This squid is big among squid species, and its skin is thick. It is clear that diamondback squid outer skin has a potential as an alternative source of collagen to bovine skin and bone. At present, collagen using aquatic materials such as skin (cod and a deep-sea fish) and scale (sea bream and anchovy) is the development stage in the related industries. Unless the problem of BSE infection in land animals is resolved aquatic materials as an alternative source of collagen will attract much attention in the cosmetic and medical fields.     

Circular-dichroism and electron-microscopy studies of human subcomponent C1q before and after limited proteolysis by pepsin.

1. A fragment of human subcomponent C1q was prepared by limited proteolysis with pepsin at 37 degrees C for 20 h, and at pH4.4, followed by gel filtration on Sephadex G-200. This fragment was shown to contain all the collagen-like features known to be present in the intact molecule [Reid (1976) Biochem. J. 155, 5-17]. 2. Circular-dichroism studies showed the presence of positive bands at 230 and 223 nm in the intact subcomponent C1q and pepsin fragment respectively, compared with a positive band at 220 nm obtained for lathyritic rat skin collagen. These bands were abolished by collagenase treatment, which suggested that there may some collagen-like triple-helical structure in subcomponent C1q and that this structure resides in the pepsin-resistant portion of the molecule. However, the 230 and 223 nm bands had a substantially lower magnitude than that obtained for the unaggregated single fibres of totally triple-helical collagen. 3. Thermal-transition temperatures obtained for subcomponent C1q, the pepsin fragment and the reduced and alkylated pepsin fragment were 48 degrees, 48 degrees and 39 degrees C respectively, compared with a value of 38 degrees C obtained for lathyritic rat skin collagen. 4. Only the unreduced pepsin fragment regained significant amounts (up to 60%) of collagen-like structure, after heat denaturation and cooling, as estimated by circular-dichroism measurements. 5. Electron-microscopy studies of subcomponent C1q and the collagen-like pepsin-resistant fragment of subcomponent C1q showed that the six peripheral globular regions of the molecule were fragmented by pepsin leaving the six collagen-like connecting strands and fibril-like central portion intact.     

Effect of oligomers of ethylene glycol on thermotropic phase transition of dipalmitoylphosphatidylcholine multilamellar vesicles.

The effect of oligomers of ethylene glycol (EG) on thermotropic phase transitions of dipalmitoylglycerophosphatidylcholine multilamellar vesicles (DPPC-MLV) were investigated. Diethylene glycol (di-EG) had a biphasic effect on transition temperature, reducing pre-transition temperature (Tp) at low concentrations but increasing main transition temperature (Tm) and extinguishing pre-transition at high concentration. Results of the X-ray diffraction method and the excimer method indicated that di-EG induced interdigitated gel phase (L beta 1 phase) in the DPPC membranes at high concentration. Phase diagram of temperature-di-EG concentration for DPPC-MLV was determined by use of X-ray diffraction and differential scanning calorimetry, which was similar to that of temperature-EG concentration. The minimum concentration of di-EG where L beta 1 phase was induced was 42%(w/v), which was larger than that of EG (30%(w/v)). On the other hand, in the presence of triethylene glycol (tri-EG), Tm and Tp increased with an increased in tri-EG concentration, as well as poly(ethylene glycol). These differences, between the effects of di-EG and those of tri-EG, might be due to the differences of their sizes.     

Phase transitions of phospholipid vesicles under osmotic stress and in the presence of ethylene glycol.

The effects of poly(ethylene glycol) (PEG) on the phase transition of phospholipid multilamellar vesicles (MLVs) were investigated by using differential scanning calorimetry (DSC). Main transition temperature (Tm) and the pre-transition temperature (Tp) of neutral phospholipid-, DMPC-1, DPPC- and DSPC-MLVs increased with an increase in PEG concentration. The subtransition temperature of DPPC-MLV also increased with an increase in PEG concentration. These results could be qualitatively explained by enhancement of the lateral packing on the basis of the osmoelastic coupling theory. The pretransition temperature increased faster than the main transition temperature did with an increase in PEG concentration. The increment of Tm depended on the hydrocarbon chain length, the shorter the hydrocarbon chain length was, the larger the increment was. The transition width in the DSC peak was broadened with an increase in PEG concentration. These three above-mentioned effects are the main differences between the effects of the osmotic stress on the phase transition of MLVs and those of hydrostatic pressure. On the other hand, ethylene glycol (EG), which is the monomer of PEG, had a biphasic effect on transition temperature of DPPC-, DSPC-, and DMPC-MLV, reducing Tm and Tp at low concentrations, but increasing Tm and extinguishing pretransition at high concentrations. This is explained by the induction of an interdigitated gel phase at high concentrations of EG, which indicates that EG can easily penetrate into the head group region of the lipid, in contrast with PEG 6K, because EG is small. Temperature-EG concentration phase diagrams for the various PC-MLVs were determined.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of various anions on the stability of the coiled coil of skeletal muscle myosin

The stability of skeletal myosin rod was studied by following the dependence of both papain digestion kinetics and helix-coil transition temperatures on the concentration of neutral salts. The rate of papain-catalyzed digestion of rod to form subfragment 2 and light meromyosin was strongly dependent on chloride concentration but essentially independent of acetate concentration up to 2.0 M. The rod exhibited a biphasic melting curve in 0.6 M NaCl, 5 mM phosphate, and 0.1 mM ethylenediaminetetraacetic acid (EDTA), pH 7.3, with transitions at 45 and 53 degrees C. In 0.6 M CH3COONa, 5 mM phosphate, and 0.1 mM EDTA, pH 7.3, the transitions occurred at 50 and 58 degrees C, respectively. Transition temperatures were obtained with a novel curve-fitting method. The effect of increasing chloride ion concentration on melting profiles was 2-fold. Below 0.6 M salt, the two transition temperatures, Tm,1 and Tm,2, depended on salt concentration such that increasing NaCl concentration caused a small stabilization of the helix while increasing acetate concentration caused the helix to become markedly more stable. Between 0.6 and 1.0 M, variation of chloride concentration had almost no effect on the thermal stability of the rod while increasing acetate concentration increased its stability considerably. Above 1.0 M NaCl, the melting profiles became broad with a third transition being observed (e.g., at 3.0 M, Tm,3 = 38 degrees C), indicating the existence of a region which has a tendency to be destabilized by chloride. The third transition was not observed at comparable concentrations of acetate. This effect of chloride was not expected on the basis of its position in the Hofmeister series.(ABSTRACT TRUNCATED AT 250 WORDS)     

Isolation and characterization of pepsin-treated type III collagen from calf skin.

Calf skin collagen was solubilized by incubating acid-extracted calf skin with pepsin at pH 2.0 and 25 degrees C, conditions that did not cause degradation of the triple helical region of collagen. Type III collagen was separated from type I collagen by differential salt precipitation at pH 7.5. The isolated type III collagen contained mainly gamma and higher molecular weight components cross-linked by reducible and/or non-reducible bonds. The isolated alpha1 (III) chains had an amino acid composition characteristic of type III collagen. Denatured but unreduced type III collagen, chromatographed on carboxymethyl-cellulose, eluted in the alpha 2 region, while after reduction and alkylation the alpha1 (III) chains eluted between the positions of alpha1 (I) and alpha2. The mid-point melting temperature temperature (tm) of type III collagen (35.1 degrees C) in a citrate buffer at pH 3.7 was somewhat lower than that of type I collagen (35.9 degrees C). Renaturation experiments at 25 degrees C showed that denatured type III collagen molecules with intact intramolecular disulfide bridges (gamma components) reform the triple helical structure of collagen much faster than reduced and carboxymethylated alpha1 (III) chains.     

Effect of propylgallate on a dipalmitoylphosphatidylglycerol and calcium mixture

Effect of propylgallate (PrG) on the thermotropic behavior of mixtures of dipalmitoylphosphatidylglycerol (DPPG) and Ca2+ was studied by means of differential scanning calorimetry (DSC). In the case of DPPG or DPPG/Ca (molar ratio, 15 : 1), the transition temperature (Tm) of the main transition and the subtransition decreased from 40 degrees C to 29 degrees C and from 29 degrees C to 20 degrees C, respectively, with an increase in the concentration of PrG. The addition of PrG to the DPPG/Ca mixture induced a shoulder on the high temperature side in the reheating scan. Neither PrG nor low concentrations of Ca2+ bind to the Lc phase of DPPG. When the molar ratio of DPPG to Ca was 1 : 1, the subtransition did not occur, that is, only the main transition (Tm = 90 degrees C) appeared. The Tm of the main transition was slightly affected by PrG. On the addition of PrG, another metastable endothermic transition peak (Tm = 78 degrees C) appeared. It is concluded that Ca2+ and PrG inhibit each other's binding.     

Discrete reduction of type I collagen thermal stability upon oxidation

The oxidation of acid-soluble calf skin collagen type I caused by metal-dependent free radical generating systems, Fe(II)/H2O2 and Cu(II)/H2O2, was found to bring down in a specific, discrete way the collagen thermal stability, as determined by microcalorimetry and scanning densitometry. Initial oxidation results in splitting of the collagen denaturational transition into two components. Along with the endotherm at 41 degrees C typical for non-oxidized collagen, a second, similarly cooperative endotherm appears at 35 degrees C and increases in enthalpy with the oxidant concentration and exposure time, while the first peak correspondingly decreases. The two transitions at 35 and 41 degrees C were registered by densitometry as stepwise increases of the collagen-specific volume. Further oxidation results in massive collagen destruction manifested as abolishment of both denaturational transitions. The two oxidative systems used produce identical effects on the collagen stability but at higher concentrations of Cu(II) in comparison to Fe(II). The discrete reduction of the protein thermal stability is accompanied by a decrease of the free amino groups, suggestive of an oxidation attack of the side chains of lysine residues. Since the denaturation temperature of collagen shifts from above to below body temperature (41 degrees C-35 degrees C) upon oxidation, it appears important to account for this effect in a context of the possible physiological implications of collagen oxidation.     

The presence of partially degraded collagen fragments in the resorbing granulation tissue in rats.

Insoluble collagen of granulation tissue produced by carrageenin injection was solubilized by pepsin treatment and purified. The pepsin-solubilized insoluble collagen contained partially degraded collagen fragments and the amounts of these small fragments of collagen were much greater in the resorbing granulation tissues than in the growing tissues, suggesting that these small fragment were formed in the course of resorption of granulation tissue, including collagen breakdown.     

Calorimetric study on a dipalmitoylphosphatidylcholine-propylgallate mixture

The effect of propylgallate (PrG, an antioxidant) on the thermotropic behavior of dipalmitoylphosphatidylcholine (DPPC) was studied by means of differential scanning calorimetry. A DPPC/PrG mixture displayed distinctive thermotropic behavior that was significantly different from that of a DPPC/cholesterol or DPPC/vitamin E mixture. Although the enthalpy of the phase transition (delta H) for DPPC decreased at a low concentration of the PrG and the transition peak became broadened, delta H increased again and the peak became sharper on the addition of more PrG. The same was observed for DPPC/methylgallate and DPPC/ethylgallate mixtures, but not for a DPPC/butylgallate mixture. On the other hand, the transition temperature (Tm) of the DPPC/gallate derivative mixtures decreased with an increase in the chain length of the acyl moiety of the gallate derivatives. The pre-transition and subtransition of the DPPC/PrG mixture were eliminated on the addition of a PrG, and Tm of the DPPC/PrG mixture approached about 26 degrees C. These results suggested that the chain length of the acyl moiety must be C1 to C3 for the unique effect of the gallate derivatives described above, and that DPPC forms a complex with PrG as a pure component.     

Heterogeneity of collagen molecules types I and II according to their resistance to proteolysis

Study of the effects of pepsin treatment on soluble collagens type I of the skin and collagens type II of the costal cartilage of healthy subjects revealed the presence of two classes of molecules differing in the stability of their three-helical structure. In collagen molecules possessing a low stability (their number may amount to 20-30%) within the temperature range of 4-30 degrees C pepsin causes a split-off of N-terminal sites with the formation of short chains, i.e., alpha 1(I), alpha 2(II), and alpha 1(II), whereas at higher temperatures (33 degrees C for collagens type I and 37 degrees C for collagens type II) a complete degradation of these molecules takes place. It was found that collagens types I and II molecules contain a high number of three-helical sites with a high susceptibility to pepsin. The putative functional role of structural heterogeneity of collagen molecules is discussed.     

Phase transition in dioctadecyldimethylammonium bromide and chloride vesicles prepared by different methods

The gel to liquid crystalline phase transition of the double-chained cationic dioctadecyldimethylammonium chloride and bromide (DODAX, X = Cl- or Br-) in aqueous vesicle dispersions prepared by non-sonication. sonication and extrusion has been investigated using high-sensitivity differential scanning calorimetry (DSC). The transition temperature (Tm) is a function of the preparation method, amphiphile concentration, vesicle curvature and nature of the counterion. DSC thermograms for DODAB and DODAC non-sonicated vesicle dispersions exhibit a single endothermic peak at Tm roughly independent of concentration up to 10 mM. Extrusion broadens the transition peak and shifts Tm downwards. Sonication, however, broadens slightly the transition peak and tends to shift Tm upwards suggesting that extrusion and sonication form vesicles with different characteristics. DODAC always exhibits higher Tm than DODAB irrespective of the preparation method. Tm changes as follows: Tm (sonicated) > or = Tm (non-sonicated) > Tm (extruded). Hysteresis of about 7 degrees C was observed for DODAB vesicle dispersions.     

Conformation, pH-induced conformational changes, and thermal unfolding of anti-p24 (HIV-1) monoclonal antibody CB4-1 and its Fab and Fc fragments

Conformation, acid-induced conformational changes and stability of the murine monoclonal antibody CB4-1 directed against the human immunodeficiency virus type 1 capsid protein p24, and its Fab and Fc fragments, were analysed by circular dichroism (CD), fluorescence, and differential scanning calorimetry (DSC) measurements. CD spectra show the characteristics expected for beta-proteins. Lowering the pH to 3.5 reduces the stability, but does not change the conformation. Between pH 3.5 and 2.0 conformational changes and the formation of new structures are indicated. Deconvolution of the bimodal DSC curves of CB4-1 reveals five 'two-state' transitions at pH 7.5. At pH 5 and below, only four transitions are found. Half transition temperatures Tm and molar enthalpy changes DeltaHm gradually decrease at pH 4 and 3.4. At pH 2.1, two low-temperature (Tm=36.9 and 44.1 degrees C) and two high-temperature (Tm=74.6 and 76.8 degrees C) transitions are identified. The Fab and Fc fragments behave similarly. Deconvolution of their monophasic DSC curves yields two 'two-state' transitions for each fragment. Tm and DeltaHm values gradually decrease at pH 4.0 and 3.4; and at pH 2.1 and 2.8 for Fab and Fc, respectively, one of the transitions is found at high temperature (Tm=67.2 and 75.9 degrees C for Fab and Fc, respectively).     

Insights into the dynamics of DMSO in phosphatidylcholine bilayers.

The solvation effects of dimethyl sulfoxide (DMSO) on the phase stability of dimyristoylphosphatidylcholine (DMPC) have been fully characterized using differential scanning calorimetry (DSC) and fluorescence spectroscopy with 1,6-diphenyl-1,3,5-hexatriene (DPH). The temperatures of the sub-, pre-, and main transitions of DMPC were found to increase linearly with increasing mole fraction of DMSO up to mole fraction X=0.13 DMSO/H(2)O. Beyond X=0.13, the pre-transition peak started to merge with the peak representing the main transition. Simultaneously, the subtransition peak began to disappear as its transition temperature also decreased. At X=0.18, with both the subtransition and pre-transition absent, the main transition between the planar gel and the liquid-crystalline phase was observed at 30.3 degrees C. Transition enthalpy values indicated that the subgel, planar gel and rippled gel phases are most stable at X=0.11, 0.16 and 0.20 DMSO/H(2)O, respectively. This demonstrates that DMSO exerts distinct effects on each respective phase and corresponding transition. Temperature-dependent fluorescence emission scans show an increase in hydration as the system proceeds from the subgel phase all the way to the liquid-crystalline phase and correlated well with the effects of DMSO on the transition temperatures of DMPC observed in our calorimetry data. Initial observations for the sub- and main transition are further confirmed by fluorescence anisotropy using DPH as a probe. The results illustrate the differences in the microviscosity of each phase and how DMSO affects the phase transitions. Ultimately, our results suggest the most likely mechanism governing the biological actions of DMSO may involve the regulation of the solvation effects of water on the phospholipid bilayer.     

Pepsin treatment of avian skin collagen. Effects on solubility, subunit composition and aggregation properties

1. Collagen was extracted from chick skin with dilute acetic acid followed by dilute acetic acid containing pepsin. 2. The solubilized collagens were purified and portions subjected to further digestion by pepsin. 3. This treatment decreased the aldehyde content but contamination by hexosamine was not diminished. 4. Pepsin treatment converted practically all the acid-soluble collagen into monomeric subunits (alpha-chains), but the pepsinsolubilized material retained a significant amount of higher subunits (beta- and gamma-chains). 5. Treatment lowered the rate of fibrillogenesis by acid-soluble collagen, but was without effect on pepsin-solubilized collagen.     

Physicochemical properties of succinylated calfskin pepsin-solubilized collagen

Some physicochemical properties of calfskin pepsin-solubilized collagen (PSC) and succinylated PSC (SPSC) were compared. The amino acid profile remained significantly unchanged. Sodium dodecylsulphate-polyacrylamide gel electrophoresis showed that subunits of SPSC migrated less than those of PSC. The denaturation temperatures of PSC and SPSC were 38.4 degrees C and 34.7 degrees C respectively. Succinylation slightly altered the triple-helical conformation of collagen, as determined by circular dichroism.