Synthetic substrates for coproporophyrinogen oxidase: mesoporphyrinogen-VI revisited

Lash, Timothy D.; Drinan, Martin A.; Zhen, Chun; Mani, Ukti N.; Jones, Marjorie A.

doi:10.1016/s0960-894x(01)80575-8

Cited by 10 publications

(10 citation statements)

References 17 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…These data provide clues about the nature of copro'gen oxidase, and this has allowed us to develop a model for the active site of this poorly understood enzyme (Scheme ) . Three regions are defined for substrate binding.…”

Section: Substrate Recognition: a Model For The Active Site Of Copro...mentioning

confidence: 99%

Normal and Abnormal Heme Biosynthesis. 1. Synthesis and Metabolism of Di- and Monocarboxylic Porphyrinogens Related to Coproporphyrinogen-III and Harderoporphyrinogen: A Model for the Active Site of Coproporphyrinogen Oxidase

et al. 1998

Self Cite

View full text Add to dashboard Cite

Coproporphyrinogen oxidase (copro'gen oxidase), which catalyses the conversion of coproporphyrinogen-III via a monovinylic intermediate to protoporphyrinogen-IX, is one of the least well understood enzymes in the heme biosynthetic pathway. To develop a model for the substrate recognition and binding recognition for this enzyme, a series of substrate analogues were prepared with two alkyl substituents on positions 13 and 17 in place of the usual propionate residues. Although the required substrate probes are porphyrinogens (hexahydroporphyrins), the corresponding porphyrin methyl esters were initialy synthesized via a,c-biladiene intermediates. These were hydrolyzed and reduced with 3% sodium amalgam to give the unstable porphyrinogens needed for the biochemical investigations. These modified structures were metabolized by avian preparations of copro'gen oxidase to give monovinylic products, but the second propionate residue was not further metabolized. In three cases, the metabolites were isolated and further characterized by proton NMR spectroscopy and mass spectrometry. When methyl or ethyl groups were placed at the 13 and 17 positions, the resulting porphyrinogens were very good substrates (although the ethyl version, mesoporphyrinogen-VI, gave slightly better results), but when propyl units were introduced metabolism was significantly inhibited and the butyl-substituted structure was only slightly transformed after long incubation periods. These results suggest the presence of an active-site lipophobic region near the catalytic site for copro'gen oxidase. The observation that the related 3-vinyl- and 3-ethylporphyrinogens with 13,17-diethyl substituents were not substrates for this enzyme confirmed the need for a second propionate residue to hold the substrate in place at the catalytic site.

show abstract

Section: Substrate Recognition: a Model For The Active Site Of Copro...mentioning

confidence: 99%

Normal and Abnormal Heme Biosynthesis. 1. Synthesis and Metabolism of Di- and Monocarboxylic Porphyrinogens Related to Coproporphyrinogen-III and Harderoporphyrinogen: A Model for the Active Site of Coproporphyrinogen Oxidase

et al. 1998

Self Cite

View full text Add to dashboard Cite

show abstract

“…Also, due to the difference in catalytic ability to perform an oxidative decarboxylation on meso'gen‐VI, it can be postulated that the two arginines are binding substrate at separate, yet fundamentally necessary, propionate moieties. The absence of propionate groups on the C and D rings of meso'gen‐VI only allows the formation of a monovinyl product (Lash et al 1994, 1999; Jones et al 2002). The ability of Arg401Ala to perform an oxidative decarboxylation on meso'gen‐VI suggests that a mutation to alanine was not detrimental to binding.…”

Section: Discussionmentioning

confidence: 99%

“…Mesoporphyrinogen‐VI (meso'gen‐VI) has been shown to be a substrate for this enzyme and undergoes only the first oxidative decarboxylation (Fig. 1) (Lash et al 1994, 1999; Jones et al 2002). The mechanism by which CPO performs catalysis is poorly understood, in part, due to the lack of knowledge about the amino acid residues involved in active site binding and catalysis.…”

mentioning

confidence: 99%

Role of aspartate 400, arginine 262, and arginine 401 in the catalytic mechanism of human coproporphyrinogen oxidase

Stephenson¹,

Stacey²,

Morgenthaler³

et al. 2007

Protein Science

View full text Add to dashboard Cite

Coproporphyrinogen oxidase (CPO) is the sixth enzyme in the heme biosynthetic pathway, catalyzing two sequential oxidative decarboxylations of propionate moieties on coproporphyrinogen-III forming protoporphyrinogen-IX through a monovinyl intermediate, harderoporphyrinogen. Site-directed mutagenesis studies were carried out on three invariant amino acids, aspartate 400, arginine 262, and arginine 401, to determine residue contribution to substrate binding and/or catalysis by human recombinant CPO. Kinetic analyses were performed on mutant enzymes incubated with three substrates, coproporphyrinogen-III, harderoporphyrinogen, or mesoporphyrinogen-VI, in order to determine catalytic ability to perform the first and/or second oxidative decarboxylation. When Asp400 was mutated to alanine no divinyl product was detected, but the production of a small amount of monovinyl product suggested the K m value for coproporphyrinogen-III did not change significantly compared to the wild-type enzyme. Upon mutation of Arg262 to alanine, CPO was again a poor catalyst for the production of a divinyl product, with a catalytic efficiency <0.01% compared to wild-type, including a 15-fold higher K m for coproporphyrinogen-III. The efficiency of divinyl product formation for mutant enzyme Arg401Ala was ;3% compared to wild-type CPO, with a threefold increase in the K m value for coproporphyrinogen-III. These data suggest Asp400, Arg262, and Arg401 are active site amino acids critical for substrate binding and/or catalysis. Possible roles for arginine 262 and 401 include coordination of carboxylate groups of coproporphyrinogen-III, while aspartate 400 may initiate deprotonation of substrate, resulting in an oxidative decarboxylation.

show abstract

“…28 A number of synthetic porphyrinogens have also been shown to be metabolized by CPO using crude enzyme preparations from chicken red cell hemolysates (CRH), including mesoporphyrinogen-VI (1a; meso'gen-VI) where the C and D ring propionate units have been replaced with ethyl groups (Scheme 3). 29,30 Meso'gen-VI and related porphyrinogens 1b-d with methyl, propyl or butyl groups at these positions are converted by CPO to the corresponding monovinyl products, but the second oxidative decarboxylation is not observed. 13,30,32 However, the specific alkyl residues at positions 13 and 17 do have an impact, as meso'gen-VI is the best substrate, followed by 1b and 1c.…”

Section: Introductionmentioning

confidence: 99%

“…29,30 Meso'gen-VI and related porphyrinogens 1b-d with methyl, propyl or butyl groups at these positions are converted by CPO to the corresponding monovinyl products, but the second oxidative decarboxylation is not observed. 13,30,32 However, the specific alkyl residues at positions 13 and 17 do have an impact, as meso'gen-VI is the best substrate, followed by 1b and 1c. Porphyrinogen 1d is a very poor substrate and only gives <10% conversion to 2d.…”

Section: Introductionmentioning

confidence: 99%

Normal and abnormal heme biosynthesis. Part 7. Synthesis and metabolism of coproporphyrinogen-III analogues with acetate or butyrate side chains on rings C and D. Development of a modified model for the active site of coproporphyrinogen oxidase

Lash

Lamm

Schaber

et al. 2011

Bioorganic & Medicinal Chemistry

Self Cite

View full text Add to dashboard Cite

Synthetic substrates for coproporophyrinogen oxidase: mesoporphyrinogen-VI revisited

Cited by 10 publications

References 17 publications

Normal and Abnormal Heme Biosynthesis. 1. Synthesis and Metabolism of Di- and Monocarboxylic Porphyrinogens Related to Coproporphyrinogen-III and Harderoporphyrinogen: A Model for the Active Site of Coproporphyrinogen Oxidase

Normal and Abnormal Heme Biosynthesis. 1. Synthesis and Metabolism of Di- and Monocarboxylic Porphyrinogens Related to Coproporphyrinogen-III and Harderoporphyrinogen: A Model for the Active Site of Coproporphyrinogen Oxidase

Role of aspartate 400, arginine 262, and arginine 401 in the catalytic mechanism of human coproporphyrinogen oxidase

Normal and abnormal heme biosynthesis. Part 7. Synthesis and metabolism of coproporphyrinogen-III analogues with acetate or butyrate side chains on rings C and D. Development of a modified model for the active site of coproporphyrinogen oxidase

Contact Info

Product

Resources

About