Pyrroline Carboxylate Reductases
Proline
Oxidoreductases Acting on CH-NH Group Donors
Pipecolic Acids
Glutamate-5-Semialdehyde Dehydrogenase
Proline Oxidase
Spin Trapping
NADP
Proline biosynthesis from L-ornithine in Clostridium sticklandii: purification of delta1-pyrroline-5-carboxylate reductase, and sequence and expression of the encoding gene, proC. (1/66)
Clostridium sticklandii utilizes combinations of amino acids for growth by Stickland reactions. Proline is an efficient electron acceptor in these reactions and is reduced to 5-aminovalerate. Proline can be partly synthesized from ornithine by the action of ornithine aminotransferase and delta1-pyrroline-5-carboxylate (PCA) reductase. Both enzymes were present in crude extracts of C. sticklandii in sufficient activity of 0.93 nkat (mg protein)(-1) and 4.3 nkat (mg protein)(-1), respectively, whereas enzymes involved in proline biosynthesis from glutamate were not detected. PCA reductase was purified to homogeneity in a three-step procedure involving ammonium sulfate precipitation, affinity chromatography with Procion Red and gel filtration on Sephadex GF200. The homogeneous enzyme was most likely an octamer of 230 kDa with a subunit size of 25 kDa as obtained by SDS-PAGE and 28.9 kDa as calculated from the sequence. Apparent Km values for PCA and NADH were 0.19 mM and 0.025 mM, respectively. The enzyme also catalysed in vitro the reverse reaction, the oxidation of proline, at alkaline pH values above 8 and higher substrate concentrations (apparent Km values: 1.55 mM for proline and 10.5 mM for NAD at pH 10.0). Studies with growing cells of C. sticklandii and [15N]proline revealed that proline is not oxidized in vivo because 15N was solely detected by HPLC-MS in 5-aminovalerate as the product of proline reduction. The proC gene encoding PCA reductase of C. sticklandii was cloned, sequenced and heterologously expressed in Escherichia coli. The enzyme exhibited high homologies to PCA reductases from different sources. Thus, C. sticklandii is able to synthesize the electron acceptor proline from ornithine (a degradation product of arginine) by action of ornithine aminotransferase and PCA reductase. (+info)A multisubunit complex of outer and inner mitochondrial membrane protein translocases stabilized in vivo by translocation intermediates. (2/66)
Translocation of nuclear encoded preproteins into the mitochondrial matrix requires the coordinated action of two translocases: one (Tom) located in the outer mitochondrial membrane and the other (Tim) located in the inner membrane. These translocases reversibly cooperate during protein import. We have previously constructed a chimeric precursor (pPGPrA) consisting of an authentic mitochondrial precursor at the N terminus (Delta(1)-pyrroline-5-carboxylate dehydrogenase, pPut) linked, through glutathione S-transferase, to protein A. When pPGPrA is expressed in yeast, it becomes irreversibly arrested during translocation across the outer and inner mitochondrial membranes. Consequently, the two membranes of mitochondria become progressively "zippered" together, forming long stretches in which they are in close contact (Schulke, N., Sepuri, N. B. V., and Pain, D. (1997) Proc. Natl. Acad. Sci. U. S. A. 94, 7314-7319). We now demonstrate that trapped PGPrA intermediates hold the import channels stably together and inhibit mitochondrial protein import and cell growth. Using IgG-Sepharose affinity chromatography of solubilized zippered membranes, we have isolated a multisubunit complex that contains all Tom and Tim components known to be essential for import of matrix-targeted proteins, namely Tom40, Tom22, Tim17, Tim23, Tim44, and matrix-localized Hsp70. Further characterization of this complex may shed light on structural features of the complete mitochondrial import machinery. (+info)A 69 bp fragment in the pyrroline-5-carboxylate reductase promoter of Arabidopsis thaliana activates minimal CaMV 35S promoter in a tissue-specific manner. (3/66)
The Arabidopsis thaliana gene that encodes pyrroline-5-carboxylate reductase (At-P5R), the last enzyme in proline biosynthesis in A. thaliana, is developmentally regulated and is highly expressed in cells that divide rapidly or undergo changes in osmotic potential. A 69 bp region (P69; -120 to -51) has previously been identified in a 5' deletion analysis of the At-P5R promoter to be necessary for the basal expression. Here, the essential role of P69 for tissue-specific expression of At-P5R is demonstrated by loss- and gain-of-function experiments. (+info)Fits, pyridoxine, and hyperprolinaemia type II. (4/66)
The rare inherited disorder hyperprolinaemia type II presents with fits in childhood, usually precipitated by infection. A diagnosis of hyperprolinaemia type II and vitamin B(6) deficiency was made in a well nourished child with fits. It is thought that pyridoxine deficiency was implicated in her fits and was the result of inactivation of the vitamin by the proline metabolite, pyrroline-5-carboxylate. (+info)Extraordinarily high density of unrelated genes showing overlapping and intraintronic transcription units. (5/66)
The cloning of pyrroline 5-carboxylate reductase from Drosophila melanogaster was accomplished by cDNA complementation of an Escherichia coli proline auxotroph. The corresponding P5cr gene is tightly clustered with three other expressed coding regions. A bidirectional promoter, an overlapping 3'UTR and an intraintronic sequence may all be found in only 4.3 kb, making this the most densely clustered region of unrelated genes in any eukaryote. (+info)The Bradyrhizobium japonicum proline biosynthesis gene proC is essential for symbiosis. (6/66)
Plant host-derived proline is proposed to serve as an energy source for rhizobia in the rhizosphere and in symbiotic root nodules. The Bradyrhizobium japonicum proC gene was isolated, and a proC mutant strain that behaved as a strict proline auxotroph in culture was constructed. The proC strain elicited undeveloped nodules on soybeans that lacked nitrogen fixation activity and plant hemoglobin. We conclude that the proC gene is essential for symbiosis and suggest that the mutant does not obtain an exogenous supply of proline in association with soybeans sufficient to satisfy its auxotrophy. (+info)Biosynthesis of proline in Pseudomonas aeruginosa. Properties of gamma-glutamyl phosphate reductase and 1-pyrroline-5-carboxylate reductase. (7/66)
gamma-Glutamyl phosphate reductase, the second enzyme of proline biosynthesis, catalyses the formation of l-glutamic acid 5-semialdehyde from gamma-glutamyl phosphate with NAD(P)H as cofactor. It was purified 150-fold from crude extracts of Pseudomonas aeruginosa PAO 1 by DEAE-cellulose chromatography and hydroxyapatite adsorption chromatography. The partially purified preparation, when assayed in the reverse of the biosynthetic direction, utilized l-1-pyrroline-5-carboxylic acid as substrate and reduced NAD(P)(+). The apparent K(m) values were: NAD(+), 0.36mm; NADP(+), 0.31mm; l-1-pyrroline-5-carboxylic acid, 4mm with NADP(+) and 8mm with NAD(+); P(i), 28mm. 3-(Phosphonoacetylamido)-l-alanine, a structural analogue of gamma-glutamyl phosphate, inhibited this enzyme competitively (K(i)=7mm). 1-Pyrroline-5-carboxylate reductase (EC 1.5.1.2), the third enzyme of proline biosynthesis, was purified 56-fold by (NH(4))(2)SO(4) fractionation, Sephadex G-150 gel filtration and DEAE-cellulose chromatography. It reduced l-1-pyrroline-5-carboxylate with NAD(P)H as a cofactor to l-proline. NADH (K(m)=0.05mm) was a better substrate than NADPH (K(m)=0.02mm). The apparent K(m) values for l-1-pyrroline-5-carboxylate were 0.12mm with NADPH and 0.09mm with NADH. The 3-acetylpyridine analogue of NAD(+) at 2mm caused 95% inhibition of the enzyme, which was also inhibited by thio-NAD(P)(+), heavy-metal ions and thiol-blocking reagents. In cells of strain PAO 1 grown on a proline-medium the activity of gamma-glutamyl kinase and gamma-glutamyl phosphate reductase was about 40% lower than in cells grown on a glutamate medium. No repressive effect of proline on 1-pyrroline-5-carboxylate reductase was observed. (+info)Multiple genes for the last step of proline biosynthesis in Bacillus subtilis. (8/66)
The complete Bacillus subtilis genome contains four genes (proG, proH, proI, and comER) with the potential to encode Delta(1)-pyrroline-5-carboxylate reductase, a proline biosynthetic enzyme. Simultaneous defects in three of these genes (proG, proH, and proI) were required to confer proline auxotrophy, indicating that the products of these genes are mostly interchangeable with respect to the last step in proline biosynthesis. (+info)Pyrroline-5-carboxylate reductase (PCR) is an enzyme that belongs to the family of oxidoreductases. Specifically, it is a part of the subclass of aldo-keto reductases. This enzyme catalyzes the chemical reaction that converts pyrroline-5-carboxylate to proline, which is an essential step in the biosynthesis of proline, an important proteinogenic amino acid.
The reaction catalyzed by PCR involves the reduction of a keto group to a hydroxyl group, and it requires the cofactor NADPH as a reducing agent. The systematic name for this enzyme is pyrroline-5-carboxylate:NADP+ oxidoreductase (proline-forming).
Deficiencies in PCR have been associated with several human diseases, including hyperprolinemia type II, a rare inherited disorder characterized by an accumulation of pyrroline-5-carboxylate and proline in body fluids.
Proline is an organic compound that is classified as a non-essential amino acid, meaning it can be produced by the human body and does not need to be obtained through the diet. It is encoded in the genetic code as the codon CCU, CCC, CCA, or CCG. Proline is a cyclic amino acid, containing an unusual secondary amine group, which forms a ring structure with its carboxyl group.
In proteins, proline acts as a structural helix breaker, disrupting the alpha-helix structure and leading to the formation of turns and bends in the protein chain. This property is important for the proper folding and function of many proteins. Proline also plays a role in the stability of collagen, a major structural protein found in connective tissues such as tendons, ligaments, and skin.
In addition to its role in protein structure, proline has been implicated in various cellular processes, including signal transduction, apoptosis, and oxidative stress response. It is also a precursor for the synthesis of other biologically important compounds such as hydroxyproline, which is found in collagen and elastin, and glutamate, an excitatory neurotransmitter in the brain.
Oxidoreductases acting on CH-NH group donors are a class of enzymes within the larger group of oxidoreductases, which are responsible for catalyzing oxidation-reduction reactions. Specifically, this subclass of enzymes acts on CH-NH group donors, where the CH-NH group is a chemical functional group consisting of a carbon atom (C) bonded to a nitrogen atom (N) via a single covalent bond.
These enzymes play a crucial role in various biological processes by transferring electrons from the CH-NH group donor to an acceptor molecule, which results in the oxidation of the donor and reduction of the acceptor. This process can lead to the formation or breakdown of chemical bonds, and plays a key role in metabolic pathways such as amino acid degradation and nitrogen fixation.
Examples of enzymes that fall within this class include:
* Amino oxidases, which catalyze the oxidative deamination of amino acids to produce alpha-keto acids, ammonia, and hydrogen peroxide.
* Transaminases, which transfer an amino group from one molecule to another, often in the process of amino acid biosynthesis or degradation.
* Amine oxidoreductases, which catalyze the oxidation of primary amines to aldehydes and secondary amines to ketones, with the concomitant reduction of molecular oxygen to hydrogen peroxide.
Pipicolic acid is not a term that refers to a specific medical condition or disease. Instead, it is a metabolite that is involved in the body's metabolic processes.
Pipicolic acid is a type of organic compound called a cyclic amino acid, which is derived from the amino acid lysine. It is produced in the liver and is excreted in urine. Pipicolic acid has been found to have various functions in the body, including regulating the metabolism of lipids and bile acids.
Abnormal levels of pipicolic acid in the body may be associated with certain medical conditions, such as liver disease or genetic disorders that affect amino acid metabolism. However, pipicolic acid is not typically used as a diagnostic marker for these conditions.
In summary, pipicolic acid is a cyclic amino acid produced in the liver and involved in various metabolic processes in the body. Abnormal levels of pipicolic acid may be associated with certain medical conditions but are not typically used as diagnostic markers.
Glutamate-5-semialdehyde dehydrogenase (G5SDH) is an enzyme that plays a crucial role in the metabolism of amino acids, specifically in the catabolic pathway of the amino acid tryptophan. G5SDH catalyzes the conversion of glutamate-5-semialdehyde to 5-aminolevulinate, which is a precursor for the synthesis of porphyrins, including heme, a component of hemoglobin.
The reaction catalyzed by G5SDH involves the oxidation of glutamate-5-semialdehyde to 5-aminolevulinate, with the concomitant reduction of NAD+ to NADH. Deficiencies in G5SDH activity can lead to neurological disorders and impaired heme synthesis, as observed in certain genetic diseases such as 2,4-dienoyl-CoA reductase deficiency and pyridoxine-dependent epilepsy.
Proline oxidase is an enzyme that catalyzes the chemical reaction of oxidizing proline to Δ^1^-pyrroline-5-carboxylate (P5C) and hydrogen peroxide (H2O2). The reaction is a part of the catabolic pathway for proline utilization in some organisms.
The systematic name for this enzyme is L-proline:oxygen oxidoreductase (deaminating, decarboxylating). It belongs to the family of oxidoreductases, specifically those acting on the CH-NH group of donors with oxygen as an acceptor. This enzyme participates in arginine and proline metabolism.
Cyclic N-oxides are a class of organic compounds that contain a cyclic structure with a nitrogen atom bonded to an oxygen atom as an N-oxide. An N-oxide is a compound in which the nitrogen atom has a positive charge and the oxygen atom has a negative charge, forming a polar covalent bond. In cyclic N-oxides, this N-O group is part of a ring structure, which can be composed of various combinations of carbon, nitrogen, and other atoms. These compounds have been studied for their potential use in pharmaceuticals, agrochemicals, and materials science.
Spin trapping is a technique used in free radical research to detect and study short-lived, reactive free radicals. It involves the use of spin trap compounds, which react with the radicals to form more stable, longer-lived radical adducts. These adducts can then be detected and analyzed using various techniques such as electron paramagnetic resonance (EPR) spectroscopy.
The spin trap compound is typically a nitrone or nitroso compound, which reacts with the free radical to form a nitroxide radical. The nitroxide radical has a characteristic EPR spectrum that can be used to identify and quantify the original free radical. This technique allows for the direct detection and measurement of free radicals in biological systems, providing valuable insights into their role in various physiological and pathological processes.
NADP (Nicotinamide Adenine Dinucleotide Phosphate) is a coenzyme that plays a crucial role as an electron carrier in various redox reactions in the human body. It exists in two forms: NADP+, which functions as an oxidizing agent and accepts electrons, and NADPH, which serves as a reducing agent and donates electrons.
NADPH is particularly important in anabolic processes, such as lipid and nucleotide synthesis, where it provides the necessary reducing equivalents to drive these reactions forward. It also plays a critical role in maintaining the cellular redox balance by participating in antioxidant defense mechanisms that neutralize harmful reactive oxygen species (ROS).
In addition, NADP is involved in various metabolic pathways, including the pentose phosphate pathway and the Calvin cycle in photosynthesis. Overall, NADP and its reduced form, NADPH, are essential molecules for maintaining proper cellular function and energy homeostasis.