The full-length, protein coding sequence for dehaloperoxidase was obtained using a reverse genetic approach and a cDNA library from marine worm Amphitrite ornata. The crystal structure of the dehaloperoxidase (DHP) was determined by the multiple isomorphous replacement method and was refined at 1.8-Å resolution. The enzyme fold is that of the globin family and, together with the amino acid sequence information, indicates that the enzyme evolved from an ancient oxygen carrier. The peroxidase activity of DHP arose mainly through changes in the positions of the proximal and distal histidines relative to those seen in globins. The structure of a complex of DHP with 4-iodophenol is also reported, and it shows that in contrast to larger heme peroxidases DHP binds organic substrates in the distal cavity. The binding is facilitated by the histidine swinging in and out of the cavity. The modeled position of the oxygen atom bound to the heme suggests that the enzymatic reaction proceeds via direct attack of the oxygen atom on the carbon atom bound to the halogen atom.Polychlorinated phenols and other polychlorinated aromatics of anthropogenic origin have been widely dispersed and constitute significant environmental problems. It is less known that bromoaromatics of biotic origin are also widespread and secreted as chemical warfare by a number of marine organisms. Dehalogenating enzymes are used as the first line of defense against these toxicants by organisms that live in such contaminated environments (1). We have recently discovered and characterized by a number of techniques (2-4) an enzyme with a novel function, dehaloperoxidase (DHP).1 DHP is isolated from Amphitrite ornata, a terebellid polychaete. This species does not produce halogenated compounds itself but usually co-habits estuarine mud flats with other polychaete worms, such as Notomastus lobatus, and hemichordata such as Saccoglossus kowalewskyi, which secrete large quantities of brominated aromatics and other halometabolites as repellents (5). The levels of DHP are very high as it represents approximately 3% of the soluble protein in crude extracts of A. ornata. The enzyme catalyzes the oxidative dehalogenation of polyhalogenated phenols in the presence of hydrogen peroxide at a rate at least 10 times faster than all known halohydrolases of bacterial origin, according to Reaction 1.The oxidative potential of hydrogen peroxide likely allows for the unusually high rate of this reaction as well as for the unique ability of DHP to dehalogenate fluorophenols. The enzyme has activity toward substrates with different numbers and positions of halogen substituents (2).The binding of oxygen and peroxide ligands and their activation are due to the presence of heme in a variety of oxygen carriers and enzymes. This is also true for DHP, which contains one heme per subunit (3) and a histidine as the proximal iron ligand (4). The propensity of peroxidases (and oxygenases, which tend to have a cysteinate proximal ligand) to cleave the oxygen-oxygen bond and form a high vale...

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Amphitrite ornata, a terebellid polychaete, inhabits marine environments that are contaminated by biogenically produced halometabolites. These halogenated organic compounds are toxic and quite diverse. To survive in this environment, A. ornata produces a novel dehaloperoxidase (DHP I) that detoxifies haloaromatic compounds. In this study we identified and characterized two dehaloperoxidase genes, designated dhpA and dhpB, from an A. ornata complementary DNA library. The deduced amino acid sequences (DHP A and DHP B) of the two dhp genes both contain 137 amino acid residues, but they differ at 5 amino acid positions. Allelic variation was observed for both genes as well. Polymerase chain reaction-restriction fragment length polymorphism assays of genomic DNA from 19 in individuals showed that each individual contains both the dhpA and the dhpB genes. Therefore, the two types of DHP are encoded by separate genes and are not alleles of a single gene. Furthermore, DHP A and DHP B may have different substrate specificities since they have amino acid differences in the active site.

Diplacus aurantiacus contains large amounts of a leaf phenolic resin, an important deterrent to a leaf-eating caterpillar, Euphydryas chalcedona. The resin can also retard water loss during drought. Furthermore, the leaf resin content differs among plants and populations. This study investigates the existence of heritable variation (h ) in resin production and tests for a genetic correlation (r ) between carbon allocation to secondary metabolites and growth rate, as well as with three other vegetative traits. Nine dam and 10 sire plants were chosen randomly at a field site and used to generate 78 full-sib families (19 half-sib families) by crossing all males to all females in a factorial design. Heritability was estimated in two ways, and genetic correlations were estimated by three methods. We found: (1) the heritability of resin production estimated by the regression of offspring on sires was significantly greater than zero (hs2=0.32, P<0.01); (2) the maternal variance in resin content was significantly greater than zero (21.3% of total phenotypic variance); (3) significant negative genetic correlation between resin content and growth rate was observed from two of three methods and was consistent with the phenotypic correlation; and (4) the cost of resin could be assessed quantitatively. The genetic cost of 1 mg in resin is equivalent to 25 mg of dry shoot-biomass growth, but the phenotypic cost is only 2.1 mg. This study indicates that carbon allocation to these secondary metabolites may respond to natural selection, and the phenotypic cost of resin production has a genetic basis in D. aurantiacus. This trade-off suggests that once selection occurs, increased phenolic resin production may result in decreased growth, or vice versa.

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