Rearrangement reactions in the biosynthesis of molybdopterin

138

124

The human proteins MOCS1A and MOCS1B catalyze the conversion of a guanosine derivative to precursor Z during molybdenum cofactor biosynthesis. MOCS1A shares homology with S-adenosylmethionine (AdoMet)-dependent radical enzymes, which catalyze the formation of protein and/or substrate radicals by reductive cleavage of AdoMet through a [4Fe-4S] cluster. Sequence analysis of MOCS1A showed two highly conserved cysteine motifs, one near the N terminus and one near the C terminus. MOCS1A was heterologously expressed in Escherichia coli and purified under aerobic and anaerobic conditions. Individual mutations of the conserved cysteines to serine revealed that all are essential for synthesis of precursor Z in vivo. The molybdenum cofactor in eukaryotic molybdoenzymes consists of a mononuclear molybdenum coordinated by the dithiolene moiety of a tricyclic pyranopterin, termed molybdopterin (MPT) 1 (1). In humans, defects in molybdenum cofactor biosynthesis lead to the pleiotropic loss of the molybdoenzymes sulfite oxidase, aldehyde oxidase, and xanthine dehydrogenase (2, 3). Affected patients usually die shortly after birth and show neurological abnormalities, such as attenuated growth of the brain, untreatable seizures, and dislocated ocular lenses (4). The first step during human molybdenum cofactor biosynthesis is catalyzed by MOCS1A and MOCS1B, leading to the synthesis of precursor Z, an oxygen-sensitive 6-alkyl pterin with a cyclic phosphate, from a guanosine derivative, most likely 5Ј-GTP (5-7).Analogous to other pteridine biosynthetic pathways, synthesis of precursor Z has been proposed to occur via a GTP cyclohydrolase-like reaction mechanism (5, 6). In contrast to these pathways, the C-8 atom of 5Ј-GTP is not released as formate but is retained and incorporated in a rearrangement reaction as the first carbon atom of the precursor Z side chain. In the second step of molybdenum cofactor biosynthesis catalyzed by MOCS3 (8) and MPT synthase (MOCS2) precursor Z is converted into MPT (9 -11). Finally molybdenum is incorporated into MPT by the multifunctional protein gephyrin (12, 13).MOCS1A contains two highly conserved cysteine motifs ( Fig. 1) proposed to be involved in iron-sulfur (FeS) cluster binding (14, 15), one is located near the N terminus (consensus sequence CX 3 CX 2 C where X denotes any amino acid), and one is near the C terminus (consensus sequence CX 2 CX 13 C). Several mutations identified in molybdenum cofactor deficiency patients are located in these conserved cysteine motifs indicating their functional importance for protein activity (2, 3). Based on sequence similarities to proteins such as biotin synthase, pyruvate formate-lyase-activating enzyme, and anaerobic ribonucleotide reductase-activating enzyme, MOCS1A has been classified as a member of the superfamily of S-adenosylmethionine (AdoMet)-dependent radical enzymes (16). In this class of enzymes, AdoMet serves as the free radical initiator and undergoes cleavage to methionine and a 5Ј-deoxyadenosyl radical that in turn propagates radical for...

Section: Discussionmentioning

confidence: 85%

“…Analogous to other pteridine biosynthetic pathways, synthesis of precursor Z has been proposed to occur via a GTP cyclohydrolase-like reaction mechanism (5,6). In contrast to these pathways, the C-8 atom of 5Ј-GTP is not released as formate but is retained and incorporated in a rearrangement reaction as the first carbon atom of the precursor Z side chain.…”

mentioning

confidence: 99%

Characterization of MOCS1A, an Oxygen-sensitive Iron-Sulfur Protein Involved in Human Molybdenum Cofactor Biosynthesis

Hänzelmann

Hernandez

Menzel

et al. 2004

138

124

“…The underlying mechanism involves a complex radical-based rearrangement reaction in which the C8 atom of the purine is inserted between the 2Ј-and 3Ј-ribose carbon atoms, thus forming the new C1Ј position in the four-carbon side chain of the pterin (28,31).…”

Section: Biosynthesis Step 1: Conversion Of Gtp To Cpmpmentioning

confidence: 99%

The Molybdenum Cofactor

Mendel

2013

244

181

The transition element molybdenum needs to be complexed by a special cofactor to gain catalytic activity. Molybdenum is bound to a unique pterin, thus forming the molybdenum cofactor (Moco), which, in different variants, is the active compound at the catalytic site of all molybdenum-containing enzymes in nature, except bacterial molybdenum nitrogenase. The biosynthesis of Moco involves the complex interaction of six proteins and is a process of four steps, which also require iron, ATP, and copper. After its synthesis, Moco is distributed, involving Mocobinding proteins. A deficiency in the biosynthesis of Moco has lethal consequences for the respective organisms.The transition element molybdenum is an essential micronutrient for microorganisms, plants, and animals (1). Surprisingly, molybdenum itself is catalytically inactive in biological systems until it is complexed by a special scaffold (2). One type of scaffold is the ubiquitous pterin-based molybdenum cofactor (Moco), 2 which, in different variants, forms part of the active centers of all molybdenum enzymes in living organisms, except one. This exception is bacterial nitrogenase, which harbors the other type of cofactor, namely the Fe-S cluster-based FeMo cofactor, which is found only once in nature (for details, compare the minireview of Hu and Ribbe (62) in this thematic series). Molybdenum belongs to the group of trace elements, i.e. the organism needs it only in minute amounts; however, unavailability of molybdenum is lethal for the organism. Molybdenum is very abundant in the oceans in the form of the molybdate anion (3). In soils, the molybdate anion is the only form of molybdenum that is available for bacteria, plants, and fungi. More than 50 enzymes are known to be molybdenumdependent. The vast majority of them are found in bacteria, whereas only seven have been identified in eukaryotes (4, 5). It is somewhat surprising that not all organisms need molybdenum. The commonly used eukaryotic model organism yeast plays no role in molybdenum research, as Saccharomyces cerevisiae does not contain either molybdenum enzymes or the Moco biosynthesis pathway. Also Schizosaccharomyces pombe does not use molybdenum, whereas Pichia pastoris needs molybdenum. Genome-wide database analyses revealed a significant number of bacteria and unicellular eukaryotes that do not need molybdenum, whereas all multicellular eukaryotes are dependent on molybdenum (6). In addition, mainly anaerobic archaea and some bacteria are molybdenum-independent, but they require tungsten for their growth (7). In the periodic table of elements, tungsten lies directly below molybdenum and features chemical properties similar to molybdenum.Molybdenum metabolism is tightly connected to Fe-S cluster synthesis in that some of the molybdenum enzymes and Moco biosynthesis itself depend on Fe-S enzymes and on a mitochondrial transporter that is known to be crucial for the maturation of cytosolic Fe-S proteins. Moreover, Moco biosynthesis recruits mechanisms previously known from Fe-S cluster synthe...

“…Genes encoding highly homologous proteins involved in MPT biosynthesis have been identified in virtually all organisms from archea to humans, indicating that the biosynthetic pathway for MPT is highly conserved (2,4). In Escherichia coli, biosynthesis of MPT begins with a guanosine derivative that is converted to the pterin intermediate, precursor Z, through the action of the moaA and moaC gene products (5,6). Subsequent conversion of precursor Z to MPT by MPT synthase involves the addition of the dithiolene sulfurs to the C-1Ј-and C-2Ј-positions of precursor Z and linearization of its cyclic phosphate.…”

Section: Molybdopterin (Mpt)mentioning

confidence: 99%

Mechanistic and Mutational Studies of Escherichia coli Molybdopterin Synthase Clarify the Final Step of Molybdopterin Biosynthesis

Wuebbens

Rajagopalan

2003

111

Biosynthesis of the molybdenum cofactor, a chelate of molybdenum or tungsten with a novel pterin, occurs in virtually all organisms including humans. In the cofactor, the metal is complexed to the unique cis-dithiolene moiety located on the pyran ring of molybdopterin. Escherichia coli molybdopterin synthase, the protein responsible for adding the dithiolene to a desulfo precursor termed precursor Z, is a dimer of dimers containing the MoaD and MoaE proteins. The sulfur used for dithiolene formation is carried in the form of a thiocarboxylate at the MoaD C terminus. Using an intein expression system for preparation of thiocarboxylated MoaD, the mechanism of the molybdopterin synthase reaction was examined.