Microeletrophoretic studies of soluble collagen

Gilbert, Isabelle

doi:10.1016/0006-3002(60)91326-3

Cited by 14 publications

(3 citation statements)

References 13 publications

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“…12, bottom) and the formation of stabilizing salt bridges at higher ionic strength may, however, lead to an increasing influence of asymmetrical ion adsorption on the net charge formation. Although preferential adsorption of chloride ions was discussed in some of the earlier studies (26,62), there is evidence for the indifferent adsorption behavior of chloride and potassium on collagen (63,64). It is, however, well known from studies on a variety of polymeric materials that preferential adsorption of hydroxide ions leads to a net negative charge of surfaces without any ionizable surface groups for pH values above pH 4 (29,65,66).…”

Section: Discussionmentioning

confidence: 99%

Electrostatic Interactions Modulate the Conformation of Collagen I

Freudenberg

Behrens²,

Welzel

et al. 2007

Biophysical Journal

104

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The pH- and electrolyte-dependent charging of collagen I fibrils was analyzed by streaming potential/streaming current experiments using the Microslit Electrokinetic Setup. Differential scanning calorimetry and circular dichroism spectroscopy were applied in similar electrolyte solutions to characterize the influence of electrostatic interactions on the conformational stability of the protein. The acid base behavior of collagen I was found to be strongly influenced by the ionic strength in KCl as well as in CaCl(2) solutions. An increase of the ionic strength with KCl from 10(-4) M to 10(-2) M shifts the isoelectric point (IEP) of the protein from pH 7.5 to 5.3. However, a similar increase of the ionic strength in CaCl(2) solutions shifts the IEP from 7.5 to above pH 9. Enhanced thermal stability with increasing ionic strength was observed by differential scanning calorimetry in both electrolyte systems. In line with this, circular dichroism spectroscopy results show an increase of the helicity with increasing ionic strength. Better screening of charged residues and the formation of salt bridges are assumed to cause the stabilization of collagen I with increasing ionic strength in both electrolyte systems. Preferential adsorption of hydroxide ions onto intrinsically uncharged sites in KCl solutions and calcium binding to negatively charged carboxylic acid moieties in CaCl(2) solutions are concluded to shift the IEP and influence the conformational stability of the protein.

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Section: Discussionmentioning

confidence: 99%

Electrostatic Interactions Modulate the Conformation of Collagen I

Freudenberg

Behrens²,

Welzel

et al. 2007

Biophysical Journal

104

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“…Microelectrophoretic studies (Gilbert 1960) have suggested that metal ions are bound t o carboxyl groups of collagen, Addition of the divalent cations lead( 11), copper (11) , calcium (11) , and magnesium (11) at constant ionic strength (0.10) increased the mobility of collagen in electrophoresis. Binding of calcium ions to citrate-soluble oxand calf-tendon collagen increases above pH 3.2 (Iron and Perkins 1962) ; this indicates that ionised carboxyl groups are responsible for the binding, Ichtyocol collagen contains about 68 mg of calcium and 35 mg of phosphate per gram (Cflimcher 1961).…”

Section: Calcium and Soft Connective Tissuementioning

confidence: 99%

“…Carboxyl groups have suggested to be responsible for the binding of calcium to collagen (Gilbert 1960, Iron and Perkins 1962). I n the present study calcium ions were found to augment the formation of insoluble collagen similarly on both the alkaline and acidic sides of the isoelectric point of collagen ; this suggests that carboxyl groups are not alone involved in the formation of insoluble collagen under the employed conditions, It is well known from the leather industry that metal ions aggregate gelatin and collagen.…”

Section: Effect Of Metal Cationsmentioning

confidence: 99%

Transformations of Rat Skin Collagen With Special Reference to the Ageing Process

Heikkinen¹

1968

Acta Physiologica Scandinavica

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Summary The formation of insoluble collagen in rat skin was studied both in vivo and in vitro in order to clarify the mechanism by which tropocollagen units are transformed into supermolecular structures, especially in relation to the ageing process. I. Suitable methods were developed for the fractionation and purification of soluble collagens (Fig. l), for the determination of the specific radioactivity of hydroxyproline in starch‐gel electrophoretic collagen fractions (Table IV, Fig. 2) and for the production of insoluble collagen in vitro (Fig. 12). II. The proportion of insoluble collagen was maximal in the skin of a newborn rat. The rapid growth period of the rat was characterized by an accumulation of collagen soluble in 0.5 M acetic acid at 4° (Fig. 3). The insoluble collagen was separated into two subfractions (designated ISC40 and ISCr). The proportion of collagen soluble in 0.5 M acetic acid upon heating at 40° (ISC40) increased with age (Figs. 4–5). ISCR could be brought into solution only after degradation of the collagen at temperatures above 60° (Fig. 5). Also the proportion of ISCR was maximal in the skin of a newborn rat (Fig. 4). The subfractions of insoluble collagen extracted by 0.5 M acetic acid at 40° from skins of 5‐day‐old rats consisted mainly of α‐ and β‐components. The proportions of β‐ and γ‐components were higher in corresponding fractions from skins of adult rats (Fig. 7). The ratio of hexmamine to hydroxyproline decreased rapidly in the insoluble collagen with advancing age (Table V). The content of free aldehyde groups was lower in the collagen of a lathyritic rat than in the collagen of a normal rat. The difference was largest in the a‐components (Table VII). The subfractions of insoluble collagen had different contents of aldehyde groups: ISC60°‐80° contained one acetaldehyde equivalent/TC less than ISC20°‐50° (Table VI). Incorporation studies indicated that insoluble collagen is formed faster in young than in adult rats (Table IX, Fig. 9). The half‐lives of 0.5 M NaC1‐soluble collagens from 6‐day‐old and 3‐month‐old rats were 27 and 73 hours, respectively (Fig. 10). The turnover of the α2‐component in 0.1 M acetic acid‐soluble collagen from a 6‐day‐old rat was faster than that of the α1‐component (Fig. 11). III. When skin slices labelled in vivo were incubated in vitro, the total and specific radioactivities of the hydroxyproline of the insoluble collagen increased during 8 hours (Table X, Fig. 12). The formation of insoluble collagen during incubation was augmented by (1) a rise in temperature of the incubation medium up to 37° (Fig. 13); (2) a decrease in the thickness of the slices to 0.3 mm (Table XI); and (3) the presence of calcium ions up to 3.0 mM concentration (Fig. 15). The 45Ca‐activity increased in the insoluble residue of skin when skin slices labelled in vivo were incubated in vitro (Table XIII). Metal cations influenced the formation of insoluble collagen in the following order of decreasing effectiveness (Table XIV and XV): Co2+ > Cu2+ > Cu+ > Au3+ = Ni2+ > Fe3+ >...

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Electric properties of macromolecules. IX. Dipole moment, polarizability, and optical anisotropy factor of collagen in solution from electric birefringence

Yoshioka

O’Konski

1966

Biopolymers

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SynopsisThe electric birefringence of collagen solutions has been measured over a wide range of field strength with the pulse technique. The soluble collagen was from rat tail tendon. The solvent used was dilute acetic acid. Very pronounced saturation of the electric birefringence was observed, permitting calculation of the optical anisotropy factor. The Kerr constant was determined by extrapolation to zero field strength. From the dependence on field strength of the birefringence, the permanent dipole moment and the anisotropy of polarizability were separately determined. The contribution of the former to the Kerr constant was found to be twice as large as that of the latter. The same conclusion was obtained from the initial slope of the rise curves of the birefringence at low fields. The permanent dipole moment was 1.5 X lo4 bebye, and the anisotropy of polarizability was about 3 X lo-" ~m .~. The magnitude of the latter indicates that the ion atmosphere polarization is important. Effects of added salt and thermal denaturation on the electric birefringence were explored.

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Microeletrophoretic studies of soluble collagen

Cited by 14 publications

References 13 publications

Electrostatic Interactions Modulate the Conformation of Collagen I

Electrostatic Interactions Modulate the Conformation of Collagen I

Transformations of Rat Skin Collagen With Special Reference to the Ageing Process

Electric properties of macromolecules. IX. Dipole moment, polarizability, and optical anisotropy factor of collagen in solution from electric birefringence

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