Fermentative pathways

The metabolic pathway for bacterial sugar fermentation proceeds through the Embden-Meyerhof-Paranas (EMP) pathway. The pathway involves many catalysed enzyme reactions which start with glucose, a six-carbon carbohydrate, and end with two moles of three carbon intermediates, pyruvate. The end pyruvate may go to lactate or be converted to acetyl CoA for the tricarboxylic acid (TCA) cycle. The fermentation pathways from pyruvate and the resulting end products are shown in Figures 9.7 and 9.8. 

Depending on the type of substrate and the microorganisms present, fermentation pathways and the products produced may vary considerably. Figure 3.3 is an example of the fermentation of sugars (only containing the elements C, H and O), illustrating that a broad range of VOCs may be formed. 

Isomerases are the largest subfamily of B 12-dependent enzymes found in bacteria, which play important roles in fermentation pathways. The only exception is methylmalonylCoA... 

J. L. Galazzo and J. E. Bailey, Fermentation pathway kinetics and metabolic flux control in suspended and immobilized Saccharomyces cerevisiae. Enzyme Microb. Technol. 12(3), 162 172 (1990). 

Clostridium acetobutylicum Escherichia [Fe] hydrogenase [NiFe] Putative cytoplasmic, ferredoxin linked H2 production during fermentation Fermentation pathway, phosphate limitation 4... 

Upon purification of the XDH from C. purinolyticum, a separate Se-labeled peak appeared eluting from a DEAE sepharose column. This second peak also appeared to contain a flavin based on UV-visible spectrum. This peak did not use xanthine as a substrate for the reduction of artificial electron acceptors (2,6 dichlor-oindophenol, DCIP), and based on this altered specificity this fraction was further studied. Subsequent purification and analysis showed the enzyme complex consisted of four subunits, and contained molybdenum, iron selenium, and FAD. The most unique property of this enzyme lies in its substrate specificity. Purine, hypoxanthine (6-OH purine), and 2-OH purine were all found to serve as reductants in the presence of DCIP, yet xanthine was not a substrate at any concentration tested. The enzyme was named purine hydroxylase to differentiate it from similar enzymes that use xanthine as a substrate. To date, this is the only enzyme in the molybdenum hydroxylase family (including aldehyde oxidoreductases) that does not hydroxylate the 8-position of the purine ring. This unique substrate specificity, coupled with the studies of Andreesen on purine fermentation pathways, suggests that xanthine is the key intermediate that is broken down in a selenium-dependent purine fermentation pathway. ... 

Figure 11.2. General features of the C02-reduction and acetate fermentation pathways showing reactions unique and common to each.

Table 11.3. Reactions unique to the acetate fermentation pathway and enzymes that catalyze them.

The carbonic anhydrase (Cam) in M. thermophila cells is elevated several fold when the energy source is shifted to acetate, suggesting a role for this enzyme in the acetate-fermentation pathway. It is proposed that Cam functions outside the cell membrane to convert CO2 to a charged species (reaction A4) thereby facilitating removal of product from the cytoplasm. Cam is the prototype of a new class (y) of carbonic anhydrases, independently evolved from the other two classes (a and P). The crystal structure of Cam reveals a novel left-handed parallel P-helix fold (Kisker et al. 1996). Apart from the histidines ligating zinc, the activesite residues of Cam have no recognizable analogs in the active sites of the a- and P-classes. Kinetic analyses establish that the enzyme has a zinc-hydroxide mechanism similar to that of Cab (Alber et al. 1999). 

In discussing the studies of Brechot et al. (24) and Peynaud et al. (25), Kunkee (I) found it odd that bacteria which ordinarily produce d or DL-lactic acid from glucose produce L-lactic acid in wine as a result of malo-lactic fermentation. Peynaud et al. (26) reported that organisms which produced only D-lactic acid from glucose produced only L-lactic acid from L-malic acid. He postulated further that the malo-lactic fermentation pathway has no free pyruvic acid as an intermediate because the optical nature of L-malic acid would be lost when it was converted to pyruvic acid since pyruvic acid has no asymmetric carbon atom. Therefore, if pyruvic acid were the intermediate, one would expect d, l, or DL-lactic acid as the end product whereas L-lactic acid is always obtained. These results lend considerable support to the hypothesis that free pyruvic... 

Other products can be produced in fermentative bacteria but the central feature of all these pathways is the strict maintenance of the oxidation-reduction balance within the fermentation system. This gives rise to another important tool in assessing fermentation pathways—a mass balance of the substrate and products. The amount of carbon, hydrogen and oxygen in the fermentation products (including cells) must correspond to the quantities in the substrate utilised. 

Barker, H. A., Amino acid degradation by anaerobic bacteria. Ann. Rev. Biochem. 50 23, 1981. A review of an important group of fermentation pathways of amino acid breakdown that occur in nature and could not be covered in this chapter. 

Parasitic stages, on the other hand, generally do not use oxygen as the final electron acceptor but use fermentative processes to obtain most of their ATP. For these stages, an uneconomical energy metabolism is not detrimental, as the host provides the nutrients. Most adult flatworms inside the final host produce end products of a fermentative carbohydrate breakdown, such as succinate, acetate, propionate and lactate. These end products are formed via malate dismutation, a fermentative pathway, which is present in all types of parasitic worms (flatworms as well as many nematodes), but which is also present in animals like freshwater snails, mussels, oysters and other marine organisms. Malate dismutation is linked to a specially... 

When plants experience anoxic conditions there is a shift in carbohydrate metabolism from an oxidative to a fermentative pathway (Fig. 1). In the absence of oxygen, ATP is generated not by the Krebs cycle but by alcoholic fermentation, i.e. glycolysis and ethanol synthesis. 

We have been studying the anaerobic response in cotton, a crop which experiences a reduction in growth rate during irrigation or waterlogging. Cotton shows a level of anaerobically inducible ADH activity comparable with that of maize, a plant which is relatively resistant to anoxia (A. Millar, unpublished data T.L. Setter, unpublished data). However, in cotton the level of the enzyme catalysing the preceding step in the fermentation pathway, PDC, is relatively low and this may lead to low rates of ethanol synthesis and hence low tolerance to anoxia. 

Type 3 fermentation, characterised by a series of reactions between mitochondrial end-products that yield branched-chain fatty acids (e.g. 2-methylvalerate, 2-methylbutyrate), occurs in Ascaris, a number of other intestinal nematodes and in some trematodes (490). However, cestodes, with the exception of Bothriocephalus scorpii, which excretes methyl-butyrate (107), have not been shown to produce branched-chain fatty acids as end-products of respiratory metabolism. Some or all of the enzymes of the TCA cycle may be present in cestodes in addition to the type 1 and type 2 fermentation pathways. The extent to which the cycle may contribute to carbohydrate metabolism in cestodes is considered below. 

Fig. 5.1. Fermentation pathways present in facultative anaerobic eukaryotes. Examples of fermentation pathways present in the cytosol and in subcellular compartments. Fermentation processes localized in hydrogenosomes (1-3) and mitochondria (4) are indicated by the shaded box. Examples of the anaerobic ATP-producing organelles shown can be found in trichomon-ads (1), chytridiomycete fungi (2), Nyctotherus ovalis (3), and parasitic helminths, bivalves and Euglena gracilis (4). CoA coenzyme A, DHAP dihydroxyacetone phosphate, Fd ferredoxin, Gly-3-P, glyceraldehyde-3-phosphate, PFO pyruvate ferredoxin oxidoreductase...

Fermentation, or the partial (02 independent) catabolism of substrates to anaerobic end products, is a second means of forming ATP. In animals, the commonest fermentative pathway is that of anaerobic glycolysis (figure 2.1). At high pH (> 8.0), the summed reaction can be written as follows ... 

It is among the most capable of invertebrate anaerobes, the helminths and the marine bivalves, that we find the best examples of alternative fermentation pathways. Many of these have been reviewed several times elsewhere, so only a brief summary will be considered here. Current concepts view the organization of anaerobic metabolism as a series of linear, and loosely linked, pathways. The most important of these, aside from classical glucose — lactate fermentation (yielding 2 moles ATP per mole glucose) are summarized by Hochachka and Somero (1984) as follows (see chapter 2) ... 

The experiment consists of incubating a small amount of 14C-labeled substrate (the pulse) with the yeast extract just long enough for each intermediate in the fermentation pathway to become labeled. The label is then chased through the pathway by the addition of excess unlabeled glucose. The chase effectively prevents any further entry of labeled glucose into the pathway. 

The 1,2-rearrangement of a substrate molecule is now recognized as a common feature of several bacterial fermentation pathways. In general, the rearrangement enables the substrate to be readily assimilated into... 

Fungicides influence the microorganism population on the crop. This may affect the fermentation pathways during processing of food, for example, the production of cheese, soy sauce, or wine. Because of these side effects, not all fungicides can be used for every purpose. Effects on the taste and fermentation must be tested before a new fungicide can be used on crops that are processed by biotechnological methods. 

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