Adsorption of cytochrome

Second, most membrane materials adsorb proteins. Worse, the adsorption is membrane-material specific and is dependent on concentration, pH, ionic strength, temperature, and so on. Adsorption has two consequences it changes the membrane pore size because solutes are adsorbed near and in membrane pores and it removes protein from the permeate by adsorption in addition to that removed by sieving. Porter (op. cit., p. 160) gives an illustrative table for adsorption of Cytochrome C on materials used for UF membranes, with values ranging from 1 to 25 percent. Because of the adsorption effects, membranes are characterized only when clean. Fouling has a dramatic effect on membrane retention, as is explained in its own section below. 

The effect of pH on the protein adsorption on CMK-3 was also investigated [152], The monolayer adsorption capacities obtained under various pH conditions are plotted in Figure 4.12, where the maximum adsorption was observed in the pH region near the isoelectric point of lysozyme (pi of about 11). Near the isoelectric point, the net charges of the lysozyme molecule are minimized and would form the most compact assembly. A similar pH effect was also seen in the adsorption of cytochrome c on CM K-3. Although the nature of the surface of mesoporous silica and... 

Fig. 1.1. (a) SPR angle shifts (A pi) showing the electrostatical adsorption of cytochrome-c on gold electrode surfaces modified with a 3-mercaptopropionic acid SAM (I) after addition of cytochrome-c to a final concentration of ca. 2.5 x 10 M in 5 mM Na-phosphate pH 7.0 (full fine) or in 5 mM Na-phosphate, 100 mM KCl pH 7.0 (dotted line) (11) after rinsing the surface with clean buffer solution, (b) Cyclic voltammograms of a monolayer of cytochrome-c adsorbed on a gold electrode modified with a 3-mercaptopropionic add SAM recorded in 5 mM Na-phosphate, pH 7.0, at 50, 100 and 200 mV s. Reproduced fix)m [214] with permission. 

A striking feature of the cytochromes c3 is their ability to modify an electrode surface in such a way as to facilitate subsequent electron transfer (21). Among the different cytochromes studied to date, only cytochrome c3 can be oxidized and reduced with high electrochemical reversibility at an electrode-solution interface without any mediator or promoter (16, 21). This reversibility contrasts with the more usual situation in which the protein absorbs on the electrode surface, thus preventing further electron transfer. Alternatively, rather slow (k° < 10 3 cm/s) heterogeneous electron transfer rates are observed. Using Miyazaki cytochrome c3 and a mercury electrode on which specific adsorption of cytochrome c3 is known to occur, the ad-... 

P-17 - Adsorption of cytochrome c onto ordered mesoporous silicates... 

P-17 - Adsorption of cytochrome c onto ordered mesoporous silicates J. Deere, E. Magner, J.G. Wall and B.K. Hodnett... 

Fig. 7.6 (A) SEIRAS spectra recorded during the adsorption of cytochrome c oxidase onto an NTA-Ni -modified Au surface via the affinity for its histidine tag introduced genetically into the protein. The bands shown are increasing in intensity with time.

It is apparent that the cytochrome c electron transfer reaction at mercury electrodes is complex and dependent on a number of parameters. The adsorption of cytochrome c at mercury is in and of itself a complicated process. The formation of the first monolayer is rapid and chemically irreversible. 

Although rapid adsorption of cytochrome c at mercury electrodes is widely accepted based on diverse experimental results, " the structure of this primary adsorption layer remains a point of controversy. If the cytochrome c... 

Adsorption of cytochrome c on bare metal electrodes has been addressed by several authors. Cotton used surface-enhanced resonance Raman... 

Haladjian et a/. studied the adsorption of cytochrome c on platinum. When a platinum electrode was first exposed to a cytochrome c solution and then washed with water the electrochemical response of Fe(II)/Fe(III) was inhibited. This result indicates that cytochrome c adsorbs strongly enough to resist removal on washing with water. 

No adsorption of cytochrome c was evident at gold in a.c. impedance measurements reported by Eddowes et at ". No detectable change in the electrode double layer capacitance was found upon addition of cytochrome c to an electrolyte solution. 

The Ag experiments are important since it is particularly feasible to detect and characterize adsorbed species by the Surface-Enhanced Raman Scattering (SERS) effect. Studies have indicated [56-58] that strong irreversible adsorption of cytochrome c occurs at Ag and that, for the less stable Fe(III) state, this is accompanied by a change in conformation. Direct evidence for this came from the spectroscopically determined reduction potential for the adsorbed protein, which showed a large negative shift, and detection of vibrations associated with non-native forms. 

Vinu A, Streb C, Hartmann M (2(X)3) Adsorption of cytochrome c on new mesoporous carbon molecular sieves. J Phys Chem B 107 8297-8299. 

A layer of phosphatidylcholine containing vinyl ferrocene immobilized at a platinum electrode has been used to make a biomembrane-like electrode [218]. This electrode provides substantial improvements in electrochemical activity of cytochrome c. Vinyl ferrocene participates in the electron transfer at this electrode as a mediator and phosphatidylcholine provides a natural biological lipid surface for adsorption of cytochrome c. 

Both ellipsometry and cyclic voltammetry were used by Szucs etal to investigate both the adsorption and the mediated and unmediated electron-transfer processes of cytochrome c on gold. The adsorption of cytochrome c was studied by ellipsometry on gold electrodes that had not been treated with surface modifiers. The measurements were carried out at 400 mV (vs. NHE), which was approximately equivalent to the open-circuit potential of the system in the absence of protein in the electrolyte. An irreversibly adsorbed layer formed that completely covered the electrode. They found the value of the calculated thickness of the adsorbed layer on bare gold to be about half that expected if cytochrome c has been in its native form, which suggested that significant unfolding occurred upon adsorption. 

Figure 4 compares the normal Raman spectrum of a solid 4-BIPY with SERS of 4-BIPY at three different electrode potentials (+0.1, -0.2, and -0.5 V) on a silver electrode. It is quite obvious that 4-BIPY remains adsorbed on the silver electrode throughout this potential range as was shown by Cotton et al. . No shift in peak positions was observed. When the silver electrode is transferred into the cytochrome c solution, 4-BIPY molecules adsorbed on the silver electrode are partially displaced by cytochrome c molecules. That is, adsorption of cytochrome c and 4-BIPY on a silver electrode are competitive and the adsorption of 4-BIPY on a silver electrode is considered to be physical in nature rather than chemisorption. Both cytochrome c and 4-BIPY molecules are located within the range of - 0.5 nm from the electrode surface, since the SERS signals of both molecules are observable at a silver electrode. 

There is a marked difference between the normal Raman signals of 2-PYS and those of a solid 2-PYS as shown in Figure 7. The SERS signals of 2-PYS are more like those of 4-PYS. The adsorption behavior of 2-PYS, however, is very different from that of 4-PYS. The adsorption of cytochrome c and 2-PYS on the silver electrode is competitive as is observed in the adsorption from the solution of cytochrome c + 4-BIPY. The SERS spectra shown in Figure 7 reveal that both 2-PYS and cytochrome c adsorb directly onto the silver electrode. 

The SERS spectrum of 2-PYCH at a silver electrode is shown in Figure 10 (curve A) and is basically the same as the normal Raman spectrum of a solid 2 PYCH. The adsorption of cytochrome c and 2-PYCH are competitive from the solution of (0.01 mM cytochrome c + 10 mM 2-PYCH) as shown in Figure 10 (curve B). Cytochrome c in the presence of 2-PYCH shows a voltammetric response visible at 0.08 V in cyclic voltammograms. In contrast to the results reported by Haladjian et al., similar voltammetric behavior of cytochrome c is observed at gold electrode in the presence of 2-PYCH. The role of 2-PYCH in the enhancement of the electrode reaction of cytochrome c is probably the same as that of 4-BIPY and 2-PYS. 

The present results clearly show that only 2-BIPY among the six pyridine derivatives used inhibits the electrode reaction of cytochrome c in the bulk of the solution. This is probably due to the formation of either stable Ag(2-BIPY)2 or a chemisorbed film of 2-BIPY on the silver electrode and these films inhibit both the adsorption of cytochrome c on the silver electrode and electron exchange between the electrode and cytochrome c in the bulk. 

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