chlorophyll-a and protoporphyrinogen

In the last common enzymatic step of tetrapyrrole biosynthesis, prior to the branching point leading to the biosynthesis of heme and chlorophyll, protoporphyrinogen IX (Protogen) is oxidised to protoporphyrin IX (Proto) by protoporphyrinogen IX oxidase (PPX). The absence of thylakoid-localised plastid terminal oxidase 2 (PTOX2) and cytochrome

Topics: Algal Proteins; Chlamydomonas reinhardtii; Chlorophyll; Cytochrome b6f Complex; Diuron; Electron Transport; Feedback, Physiological; Gene Expression Regulation, Plant; Herbicides; Oxidation-Reduction; Photosynthesis; Plant Proteins; Plastids; Plastoquinone; Protoporphyrinogen Oxidase; Protoporphyrins

Two chlorophyll-deficient mutants of Chlamydomonas reinhardtii, chl1 and brs-1, are light sensitive and, when grown heterotrophically in the dark, accumulate protoporphyrin IX and exhibit yellow/orange pigmentation. The lesions in both mutants were mapped to the gene (CHLH) for the plastid-localized H subunit of the heterotrimeric magnesium chelatase that catalyzes the insertion of magnesium into protoporphyrin IX. The genetic defects in the mutants could be assigned to +1 frameshift mutations in exon 9 (chl1) and exon 10 (brs-1) of the CHLH gene. In both mutants, the H subunit of magnesium chelatase was undetectable, but, as shown for chl1, the steady-state levels of the I and D subunits were unaltered in comparison to wild type. The CHLH gene exhibits marked light inducibility: levels of both the mRNA and the protein product are strongly increased when cultures are shifted from from the dark into the light, suggesting that this protein may play a crucial role in the light regulation of chlorophyll biosynthesis.

Topics: Amino Acid Sequence; Aminolevulinic Acid; Animals; Bacterial Proteins; Blotting, Southern; Blotting, Western; Carboxypeptidases; Chlamydomonas; Chlorophyll; Chloroplasts; Chromosomal Proteins, Non-Histone; Cloning, Molecular; DNA Primers; Frameshift Mutation; Fungal Proteins; Genetic Complementation Test; Lyases; Methyltransferases; Molecular Sequence Data; Polymerase Chain Reaction; Porphyrins; Protoporphyrins; RNA, Messenger; Saccharomyces cerevisiae Proteins

Oleic acid stimulates enzymatic protoporphyrinogen oxidation by extracts of barley mitochondria and etioplasts. Greater stimulation occurred with Triton X-100 extracts which had been passed over a Sephacryl S-200 column than with crude Triton extracts, suggesting that purification may have removed a lipid factor required for optimal enzymatic activity. Palmitic acid, various phospholipids and detergents, or esters and alcohols of oleic acid did not substitute for free oleic acid. Linoleic acid caused a greater stimulation of protoporphyrinogen oxidation in both crude and purified barley organelle extracts and also caused a slow chemical oxidation of protoporphyrinogen. The stimulating effect of unsaturated fatty acids on enzymatic protoporphyrinogen oxidation may indicate a lipid requirement for this membrane bound enzyme or may also indicate involvement of unsaturated lipid oxidation in plant protoporphyrinogen oxidation.

Topics: Chlorophyll; Fatty Acids, Unsaturated; Heme; Hordeum; Linoleic Acid; Linoleic Acids; Mitochondria; Oleic Acid; Oleic Acids; Organoids; Oxidation-Reduction; Palmitic Acid; Palmitic Acids; Plants; Porphyrins; Protoporphyrins

High rates of oxidation of protoporphyrinogen to protoporphyrin were demonstrable in etioplasts, chloroplasts, and mitochondria from young barley shoots. Much lower rates were observed in chloroplasts from older barley or mature spinach, in mitochondria from potatoes or rat liver, and in membranes from the bacteria Escherichia coli and Rhodopseudomonas spheroides. The presence of high activity in cells capable of rapid synthesis of large amounts of chlorophyll suggests a role for this activity in chlorophyll synthesis. Characteristics of the plant protoporphyrinogen-oxidizing activity were compared to the activity in rat liver mitochondria. The activity in spinach chloroplasts exhibited a pH optimum of 7, which was lower than that of the mammalian enzyme. The plant activity was more sensitive to inhibition by glutathione or excess detergent, and was more readily inactivated at room temperature. The plant activity exhibited less specificity toward porphyrinogen substrates, oxidizing mesoporphyrinogen as rapidly as protoporphyrinogen. The mammalian enzyme oxidized mesoporphyrinogen slowly, and neither system oxidized coproporphyrinogen or uroporphyrinogen. Both the plant and the mammalian activity were bound to organelle membranes, but could be extracted with detergents. In contrast, activity from membranes of the bacteria E. coli and R. spheroides was inactivated by detergent treatment. The plant extracts could be fractionated with ammonium sulfate and retained activity after dialysis or Sephadex G-25 treatment, suggesting no readily dissociable cofactor. The activity extracted from spinach chloroplasts was mostly inactivated by trypsin digestion, which was additional evidence for the protein nature of the plant activity.

Topics: Animals; Chlorophyll; Chloroplasts; Detergents; Heme; Hordeum; Mitochondria; Mitochondria, Liver; Organoids; Oxidation-Reduction; Oxidoreductases; Plants; Porphyrins; Protoporphyrins; Rats; Substrate Specificity

Topics: Chlorophyll; Chloroplasts; Heme; Hordeum; Mitochondria; Oxidation-Reduction; Porphyrins; Protoporphyrins