RESUMEN
In plants, phosphatidylcholine is the major phospholipid in extra-plastid membranes and is synthesised mainly by the CDP-choline pathway. Evidence from studies in animals, as well as in plants, suggests that the intermediate step catalysed by cholinephosphate cytidylyltransferase (CPCT) has a major control in carbon flux to this lipid. We have isolated a full-length CPCT cDNA (designated PCT2) from Pisum sativum cv. Feltham First using an Arabidopsis probe and the polymerase chain reaction (PCR). The deduced amino acid of PCT2 is 48%, 43% and 76% identical to the rat, yeast and Brassica napus amino acid sequences, respectively. Expression of the CPCT protein in Escherichia coli confirmed the activity of the enzyme. Expression of the PCT2 mRNA in pea roots and stems was increased by treatment with 0.1 microM indole-3-acetic acid.
Asunto(s)
Citidililtransferasa de Colina-Fosfato/genética , Pisum sativum/genética , Proteínas de Plantas/genética , Secuencia de Aminoácidos , Citidililtransferasa de Colina-Fosfato/biosíntesis , ADN Complementario/genética , Regulación de la Expresión Génica de las Plantas , Ácidos Indolacéticos/farmacología , Datos de Secuencia Molecular , Pisum sativum/enzimología , Fosfatidilcolinas/biosíntesis , Reguladores del Crecimiento de las Plantas/farmacología , Proteínas de Plantas/biosíntesis , Raíces de Plantas/enzimología , Proteínas Recombinantes/biosíntesis , Homología de Secuencia de AminoácidoRESUMEN
Pseudomonas putida morphine dehydrogenase is shown to be closely homologous to 18 proteins, defining a superfamily within which morphine dehydrogenase particularly resembles two bacterial, 2,5-dioxo-D-gluconic acid reductases, and two eukaryotic proteins of unknown functions. Relationships within the superfamily are extensive and complex. Residue identities between protein pairs range from 29-90%. Three subgroups are proposed. Nevertheless, on the basis of residue conservations/exchanges it is suggested that the nicotinamide coenzyme binding and substrate reduction occur in all the enzymes by broadly analogous mechanisms, among which some probable differences are identified.
Asunto(s)
Oxidorreductasas de Alcohol/metabolismo , Pseudomonas putida/enzimología , Oxidorreductasas de Alcohol/química , Oxidorreductasas de Alcohol/clasificación , Aldehído Reductasa/química , Aldehído Reductasa/metabolismo , Secuencia de Aminoácidos , Secuencia de Bases , Sitios de Unión , Coenzimas/metabolismo , ADN Complementario , Humanos , Hidrógeno/química , Datos de Secuencia Molecular , Oxidorreductasas de Alcohol Dependientes de NAD (+) y NADP (+) , Prostaglandina-Endoperóxido Sintasas/química , Prostaglandina-Endoperóxido Sintasas/metabolismo , Conformación Proteica , Alineación de SecuenciaRESUMEN
The nucleotide sequence of 3.2 kbp of pea chloroplast DNA located upstream from the petA gene for cytochrome f, and previously reported to contain the gene for a photosystem I polypeptide, has been determined. Three open reading frames of 587, 40 and 157 codons have been identified. Orf40 encodes a highly conserved, hydrophobic, membrane-spanning polypeptide, and is identified as the gene psaI for the 4 kDa subunit of photosystem I. Orf587 is an extended version of the gene zfpA previously identified as encoding a conserved putative zinc-finger protein. The product of orf587 shows extensive homology to an unidentified open reading frame cotranscribed with a gene for folate metabolism in Escherichia coli and local homology to a region of the beta subunit of rat mitochondrial propionyl-CoA carboxylase. It is suggested that the product of orf587 is an enzyme of C1 metabolism and is unlikely to be a regulatory DNA-binding protein. Orf157 potentially encodes an unidentified basic protein, but the protein sequence is not conserved in other plants.
Asunto(s)
Cloroplastos , Fabaceae/genética , Proteínas del Complejo del Centro de Reacción Fotosintética/genética , Plantas Medicinales , Secuencia de Aminoácidos , Secuencia de Bases , Evolución Biológica , Western Blotting , ADN/genética , Datos de Secuencia Molecular , Sistemas de Lectura Abierta , Complejo de Proteína del Fotosistema I , Mapeo Restrictivo , Homología de Secuencia de Ácido NucleicoRESUMEN
The nucleotide sequence of a 1 kbp region of pea chloroplast DNA upstream from the gene petA encoding apocytochrome f has been determined. An open reading frame of 231 codons (ORF231) encoding a putative membrane-spanning polypeptide is separated by 205 bp from the coding region of petA. The open reading frame is homologous to open reading frames located in a similar position with respect to petA in chloroplast DNA from Marchantia polymorpha, tobacco, rice, wheat and Vicia faba. The sequence around a conserved histidine residue in a putative membrane-spanning region of the polypeptide resembles sequences present in cytochrome b from chromaffin granules and neutrophil membranes, suggesting that the open reading frame may encode a haem-binding polypeptide, possibly a b-type cytochrome. Northern hybridisation analysis indicates the presence in pea chloroplasts of a complex pattern of transcripts containing ORF231. Large transcripts of 5.5 kb, 4.3 kb, 3.4 kb and 2.7 kb encode both ORF231 and apocytochrome f, indicating that ORF231 and petA are co-transcribed.
Asunto(s)
Apoproteínas/genética , Proteínas Portadoras/genética , Cloroplastos/metabolismo , Citocromos/genética , Fabaceae/genética , Genes de Plantas , Hemoproteínas , Proteínas de la Membrana/genética , Proteínas de Plantas/genética , Plantas Medicinales , Secuencia de Aminoácidos , Apoproteínas/biosíntesis , Secuencia de Bases , Proteínas Portadoras/biosíntesis , Grupo Citocromo b/genética , Citocromos/biosíntesis , Citocromos f , Herencia Extracromosómica , Regulación de la Expresión Génica , Proteínas de Unión al Hemo , Proteínas de la Membrana/biosíntesis , Datos de Secuencia Molecular , Sistemas de Lectura Abierta , Proteínas de Plantas/biosíntesis , Plantas/genética , Conformación Proteica , ARN Mensajero/biosíntesis , Homología de Secuencia de Ácido Nucleico , Especificidad de la Especie , Transcripción GenéticaRESUMEN
The genes encoding the 9 kDa and 4 kDa polypeptides of cytochrome b-559 have been located in pea chloroplast DNA by coupled transcription-translation of cloned restriction fragments of chloroplast DNA in a cell-free extract of Escherichia coli and by nucleotide sequence analysis. The genes (psbE and psbF) are located approximately 1.0 kbp downstream of the gene for cytochrome f and are transcribed in the opposite direction, similar to the arrangement in the chloroplast genomes of other higher plants. Nucleotide sequence analysis of this region revealed four open reading frames encoding hydrophobic proteins of 83 (psbE), 39 (psbF), 38 and 40 amino acid residues, which are co-transcribed as a single major RNA of 1.1 kb. The 5' and 3' ends of this RNA have been located by primer extension and S1 nuclease mapping. The 5' end of the RNA is located 140 bp upstream of the initiating ATG codon of psbE and is preceded by typical chloroplast promoter sequences. The 3' end of the RNA is located approximately 515 bp downstream of the TAA stop codon of psbF close to a stable stem-loop structure.