Marine environment enzymes

Compared with tar, which has a relatively short lifetime in the marine environment, the residence times of plastic, glass and non-corrodible metallic debris are indefinite. Most plastic articles are fabricated from polyethylene, polystyrene or polyvinyl chloride. With molecular weights ranging to over 500,000, the only chemical reactivity of these polymers is derived from any residual unsaturation and, therefore, they are essentially inert chemically and photochemically. Further, since indigenous microflora lack the enzyme systems necessary to degrade most of these polymers, articles manufactured from them are highly resistant or virtually immune to biodegradation. That is, the properties that render plastics so durable... 

Obviously we must obtain considerably more information concerning the causative nature of this interesting enzyme response before any of the species we are currently investigating can be routinely used to monitor the marine environment for pollutants. 

The marine environment acts as a sink for a large proportion of polyaromatic hydrocarbons (PAH) and these compounds have become a major area of interest in aquatic toxicology. Mixed function oxidases (MFO) are a class of microsomal enzymes involved in oxidative transformation, the primary biochemical process in hydrocarbon detoxification as well as mutagen-carcinogen activation (1,2). The reactions carried out by these enzymes are mediated by multiple forms of cytochrome P-450 which controls the substrate specificity of the system (3). One class of MFO, the aromatic hydrocarbon hydroxylases (AHH), has received considerable attention in relation to their role in hydrocarbon hydroxylation. AHH are found in various species of fish (4) and although limited data is available it appears that these enzymes may be present in a variety of aquatic animals (5,6,7,8). 

Owing to the immense diversity of animals and microbes in the marine environments, and their almost untouched capacity to produce natural products, the importance for marine pharmaceutical and nutra-ceutical industries was realized on a broad basis by the scientific communities recently. This has strengthened worldwide research activities on the exploration of bioactive compounds from marine organisms for pharmaceutical use, as well as the development of other valuable products, such as enzymes, nutraceuticals, and cosmeceuticals. 

Helmke, E., and H. Weyland. 1991. Effect of temperature on extracellular enzymes occurring in permanently cold marine environments. Kieler Meeresforschungen Sonderheft 8 198-204. 

Modification of Proteins with Proteolytic Enzymes from the Marine Environment... 

Enzymatic assays can be applied in the marine environment to provide indirect information on dissolved compounds that are available to fuel bacterial production. Approaches that have been commonly appHed include measuring hydrolytic enzyme activities in seawater and monitoring degradation rates of model compounds. Protein hydrolysis in seawater is rapid as expressed by model protein studies (e.g., Nunn et al., 2003 Pantoja and Lee, 1999). This rapid and selective removal of dissolved proteins explains the relatively minor contribution from proteins to the accumulating DOM reservoir even though proteins are by far the most abundant intracellular biochemical. In an elegant study, Nunn and coworkers (2003) used matrix assisted laser desorption/ionization (MALDI) time of flight (TOP) mass... 

Hoppe, H. G., Arnosti, C., and Herndl, G. (2002). Ecological significance of bacterial enzymes in the marine environment. In Enzymes in the Environment Activity, Ecology and Applications (Bums, R., and Dick, R., eds.). Marcel Dekker, New York. pp. 73-107. 

It is also possible to use antibody based approaches to assay the accumulation of enzymes involved in denitrification pathways in environmental samples. Ward and Cockroft (1993) developed antisera to nitrite reductase from a marine Pseudomonas stutzeri isolate, and assayed sediment, water column and microbial mat samples. The antibody identified nitrite reductase in mat samples and was more specific than a DNA probe designed against the same species (Ward and Cockroft, 1993). JVIore recently, other strains of denitrifiers have been isolated from both sediment and water column samples. Sequence information regarding the denitrifying genes should prove invaluable for future refinement of both sequence based approaches and the development of additional antibodies to denitrifiers important in the marine environment. 

Enzyme-mediated reactions are responsible for most of the transformations of nitrogen (N) that occur in the marine environment, thus measurements of enzymes have figured prominently in oceanographic work for some time, especially in terms of N assimilation (see Falkowski, 1983). 

N is often limiting in the marine environment. Further, many enzymes are sensitive to cellular substrate concentrations rather than extracellular concentrations and it is difficult to measure the relevant intracellular metabohte pools. In vitro assays may affect the conformation of enzymes and the degree to which they are modified. For example, allosteric effects (see Section 1.3.3) may be modified under in vitro conditions. Many enzymes undergo posttranslational regulation wherein enzyme activity is affected by binding of activator/inactivator proteins and covalent modification of the enzyme (e.g., adenylylation, phosphorylation or carbamylation) (Ottaway, 1988). When there is posttranslational modification of enzymes, enzyme activity measured in assays may be unrelated to in vivo activity (see Section 2.2.1) and there are few ways to determine the extent of enzyme modification in nature. 

Although inorganic N has traditionally been thought to be the primary N source for marine autotrophs, it is now apparent that this is not always true (see Chapter 7 by Mulholland and Lomas, this volume) and that organic N can be a major N source in many marine environments. With that in mind, enzymes involved in the hydrolysis or mobihzation of organic N are examined. 

There are fewer measurements of endopeptidase activity in aquatic samples (see below). In contrast, there have been numerous measurements of LAP, an exopeptidase, in freshwater and marine environments using artificial fluorogenic substrates, such as Leu-AMC (Martinez et al, 1996 Rosso and Azam, 1987 Sinsabaugh et al, 1997 Stoecker and Gustafson, 2003 Stoecker et al, 2005). LAP hydrolyzes a broad spectrum of substrates with a free amino group, but has a preference for N-terminal leucine and related amino acids (Mahler and Cordes, 1966). Measuring LAP may underestimate peptide utilization not only because dipeptides are directly incorporated (see below), but also because endopeptidases are not the only enzymes that hydrolyze protein-like material. 

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