Ascorbate oxidase cytochrome oxidases

The reader will also find in the Index certain broad classifications of components, like oxidases and free radicals. These and similar examples in the Index are not there to confuse the reader, as many of the individual components in the broad classifications have specific CAS numbers. Generally, the references associated with these classes of components (found within the chapters noted in the Index) will provide the reader with information of a common nature. In nearly all cases, individual components such as ascorbate oxidase, choline oxidase, cytochrome oxidase, and glycolate oxidase follow after the broadly classified component, oxidase. Likewise, specific free radicals such as methyl-acyl radical, ethyl-acyl radical, and propyl-acyl radical 2 isomers may be found in the Index. For some components in the Index, several partially identified isomers exist, their number noted, and included in the total number of components identified in tobacco and/or smoke. 

In addition to its infiuence on blood formation copper has been shown t( be concerned in many other functions within the body. A number of oxi dases which play a part in the oxidation-reduction processes of the cell are known to be copper-protein compounds. These include the polypheAol oxidases of mushrooms and potatoes and ascorbic acid oxidase. Cytochrome oxidase is also usually considered a copper compound, although this is not certain. It is generally believed that the copper ion is the reversibly attached prosthetic group of the enzyme whose activity is associated with a valency change of the copper. An adequate intake of copper has been shown to be essential for the formation and maintenance of normal cytochrome oxidase and catalase activity of certain tissues in the rat. ... 

Nitric oxide reductase (P) Nitrous oxide reductase (P) Ascorbate oxidase (P) Cytochrome oxidase (PM) Copper ATPase pumps (PM)... 

In addition to binding to cytochrome c oxidase, cyanide inhibits catalase, peroxidase, methemoglobin, hydroxocobalamin, phosphatase, tyrosinase, ascorbic acid oxidase, xanthine oxidase, and succinic dehydrogenase activities. These reactions may make contributions to the signs of cyanide toxicity (Ardelt et al. 1989 Rieders 1971). Signs of cyanide intoxication include an initial hyperpnea followed by dyspnea and then convulsions (Rieders 1971 Way 1984). These effects are due to initial stimulation of carotid and aortic bodies and effects on the central nervous system. Death is caused by respiratory collapse resulting from central nervous system toxicity. 

Freebaim noted a decrease in oxygen uptake of plant and bovine liver mitochondria that was reversible by glutathione and ascorbic acid. The activity of some mitochondrial enzymes, including succinic dehydrogenase and cytochrome oxidase, has been found to be susceptible to ozone. 

In the oxidation of ascorbate by Oj catalysed by ascorbate oxidase, the formation of the monodehydroascorbate free radical was demonstrated by EPR spectroscopy in a flow cell. A steady state was usually reached within 50 ms. The production of the free radical was also followed by the reduction of Fe(in)-cytochrome c. Thus the oxidation of ascorbate occurs in a one-electron step The formation of the monodehydroascorbate free radical was also measured directly by spectrophotometry at 360 nm, where the free radical shows an absorption maximum... 

Copper has an essential role in a number of enzymes, notably those involved in the catalysis of electron transfer and in the transport of dioxygen and the catalysis of its reactions. The latter topic is discussed in Section 62.1.12. Hemocyanin, the copper-containing dioxygen carrier, is considered in Section 62.1.12.3.8, while the important role of copper in oxidases is exemplified in cytochrome oxidase, the terminal member of the mitochondrial electron-transfer chain (62.1.12.4), the multicopper blue oxidases such as laccase, ascorbate oxidase and ceruloplasmin (62.1.12.6) and the non-blue oxidases (62.12.7). Copper is also involved in the Cu/Zn-superoxide dismutases (62.1.12.8.1) and a number of hydroxylases, such as tyrosinase (62.1.12.11.2) and dopamine-jS-hydroxylase (62.1.12.11.3). Tyrosinase and hemocyanin have similar binuclear copper centres. 

In the discussion of the biochemistry of copper in Section 62.1.8 it was noted that three types of copper exist in copper enzymes. These are type 1 ( blue copper centres) type 2 ( normal copper centres) and type 3 (which occur as coupled pairs). All three classes are present in the blue copper oxidases laccase, ascorbate oxidase and ceruloplasmin. Laccase contains four copper ions per molecule, and the other two contain eight copper ions per molecule. In all cases oxidation of substrate is linked to the four-electron reduction of dioxygen to water. Unlike cytochrome oxidase, these are water-soluble enzymes, and so are convenient systems for studying the problems of multielectron redox reactions. The type 3 pair of copper centres constitutes the 02-reducing sites in these enzymes, and provides a two-electron pathway to peroxide, bypassing the formation of superoxide. Laccase also contains one type 1 and one type 2 centre. While ascorbate oxidase contains eight copper ions per molecule, so far ESR and analysis data have led to the identification of type 1 (two), type 2 (two) and type 3 (four) copper centres. 

There are other modes of inactivation of pressor amines that probably are not highly significant factors from our present standpoint. These include the effect of the cytochrome C-cytochrome oxidase oxidation of catechol derivatives to the corresponding ortho-quinone (41, 103), the oxidation of the phenolic nucleus by the ascorbic-dehydroascorbic acid system (18), and the deamination of pressor amines in the presence of aldehydes (120, 121,152). One may refer to the reviews by Hartung (89) and by Beyer (23) for discussions of these systems. 

Blue Multicopper Oxidases. These include laccases, ascorbate oxidase, and ceruloplasmin [22,61], which along with cytochrome c oxidase (CcO with Fe and Cu) can couple the one-electron oxidation of substrates (e.g., ascorbate, diamines, monophenols Fe2+ for ceruloplasmin cytochrome c, for CcO) to the full reduction of dioxygen to water (i.e., 02 + 4c + H+ —> 2H20). 

Figure 5.1 Schematic representations of selected active sites of the copper proteins plastocyanin [56] (type 1, a) galactose oxidase [57] (type 2, b) oxy hemocyanin [58] (type 3, c) ascorbate oxidase [10] (type 4, or multicopper site, d) nitrous oxide reductase [59] (CuA site, e) cytochrome c oxidase [15]...

Oxidation galactose oxidase, amine oxidase, ascorbate oxidase, laccase, cytochrome c oxidase... 

Multicopper blue oxidases are synthesized as a single polypeptide chain, which is composed of three BCB domains in the case of laccases (LC) and ascorbate oxidases (AO) and six such domains in ceruloplasmin (CP) and hephaestin (HP). Structurally they are arranged in a triangular manner. These enzymes, along with heme-copper oxidases (cytochrome c oxidases and quinol-oxidases) and a cyanide-resistant alternative oxidase found in mitochondria of plants and fungi, are the only known enzymes capable of catalyzing four-electron reduction of dioxygen to water. In the... 

Ascorbate oxidase, laccase, and ceruloplasmin form the group of blue oxidases. These are multicopper enzymes catalyzing the four-electron reduction of dioxygen to water with concomitant one-electron oxidation of the substrate (3), which is very similar to the reaction performed by cytochrome c oxidase. All three enzymes have been known for many years, and an overwhelming number of papers have appeared since their discovery dealing with the different aspects of these enzymes. 

The group of small plant proteins, azurin, stellacyanin, and plasto-cyanin, appear to be electron transfer proteins. They are listed because they share a type of copper site with the intensely blue representatives of the first class like laccase and ascorbate oxidase. The evidence that they participate in plant electron transfer chains remains circumstantial. Azurins, for example, purify along with well-known respiratory chain proteins like cytochrome C. A good deal of evidence exists, however, that plastocyanin is important in the photoreduction of NADP see below). 

A cytochrome b (M, 30000) from S. acidocaldarius (DSM 639) contains a single copper atom and has an a band located at 558 mn when determined at room temperature and a split a band (553 and 562 nm) when measured at liquid-nitrogen temperatures. This cytochrome reacts with CO and may function as an o-type cytochrome oxidase. Solubilization results in the loss of catalytic activity with either TMPD-ascorbate or caldariella quinone as a consequence of either the loss of lipids or polypeptides that constituted a integral part of the oxidase system. 

S. acidocaldarius (strain 7) contains a cyanide-sensitive cytochrome oxidase [24], The purified cytochrome (M, 150000) is composed of three subunits (M, 37000, 23 000, and 14000). Difference spectra following reduction with dithionite show a Soret band at 441 nm and a maximum at 603 nm characteristic of aa3-type cytochromes. In addition, there is a band at 558 nm whose connection to the oxidase is not clear. This oxidase is stimulated by cholate, but unlike the oxidase from the DSM 639 strain it is inhibited by low concentrations of cyanide (pM as opposed to mM) and oxidizes horse-heart cytochrome c, TMPD-ascorbate, and caldariella quinol. The rates of oxidation (pmol/min/mg protein) for cytochrome c, TMPD-ascorbate, and quinol are 63, 6.1, and 0.2, respectively. Another cytochrome oxidase that has an absorption maximum at 602 nm, oxidizes caldariella quinol, but does not oxidize cytochrome c, is also present in strain 7 so that the terminal portion of the electron transport system in S. acidocaldarius consists of at least three oxidases. It is suggested [8] that the presence of three oxidases in 5. acidocaldarius is unlikely and that the cyanide-sensitive oxidase was isolated from a different species, namely S. solfataricus. There is little taxonomic information in this assertion to judge whether strain 7 and DSM 639 are indeed different species. However, based on growth conditions reported by the investigators [12,28], which are unique for S. acidocaldarius and S. solfataricus [ 22, there is no reason to suspect that these organisms are different species. 

Rice bran contains active enzymes (30). Germ and the outer layers of the caryopsis have higher enzyme activities. Some enzymes that are present include a-amylase, p-amylase, ascorbic acid oxidase, catalase, cytochrome oxidase, dehydrogenase, deoxyribonuclease, esterase, flavin oxidase, a and p-glycosidase, invertase, lecithi-nase, lipase, lipoxygenase, pectinase, peroxidase, phosphatase, phytase, proteinase, and succinate dehydrogenase. 

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