The structural basis for seryl-adenylate and Ap4A synthesis by seryl-tRNA synthetase

Belrhali, Hassan; Yaremchuk, A.; Tukalo, M. A.; Berthet-Colominas, C.; Rasmussen, Bjarne; Bösecke, P.; Diat, Olivier; Cusack, Stephen

doi:10.1016/s0969-2126(01)00166-6

Cited by 87 publications

(96 citation statements)

References 40 publications

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“…Finally, the side chain of Asp-76 moves 0.9 Å to hydrogen bond with the adenine base 6-amino group, replacing a bound water molecule in the apo-enzyme structure. These changes are consistent with the induced-fit mechanism of ATP binding, known for class II AARSs (18,19,(22)(23)(24)(25).…”

Section: Resultssupporting

confidence: 88%

Breaking sieve for steric exclusion of a noncognate amino acid from active site of a tRNA synthetase

Swairjo

Schimmel²

2005

Proc. Natl. Acad. Sci. U.S.A.

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The genetic code is fixed in aminoacylation reactions catalyzed by aminoacyl-tRNA synthetases. Amino acid discrimination occurs at two sites: one for amino acid activation and aminoacylation and one for editing misactivated amino acids. Although the active site sieves out bulkier amino acids, misactivation occurs with substrates whose side chains are smaller than the cognate one. Paradoxically, although alanyl-tRNA synthetase activates glycine as well as alanine, the sterically larger (than alanine) serine is also misactivated. Here, we report crystal structures of an active fragment of Aquifex aeolicus alanyl-tRNA synthetase complexed, separately, with Mg 2؉ -ATP, alanine, glycine, and serine. Ala and Gly are bound in similar orientations in a side-chain-accommodating pocket, where ␣-amino and carboxyl groups are stabilized by salt bridges, and the carboxyl by an H-bond from the side chain NH2 of Asn-194. In contrast, whereas the same two salt bridges stabilize bound Ser, H-bonding of the highly conserved (among class II tRNA synthetases) Asn-194 side chain NH 2 to the Ser OH, instead of to the carboxyl, forces pocket expansion. Significantly, in the Mg 2؉ -ATP complex, Asn-194 coordinates a Mg 2؉ -␣-phosphate bridge. Thus, the sieve for Ser exclusion is broken because of selective pressure to retain Asn-194 for Mg 2؉ -ATP and Ala binding.alanyl-tRNA synthetase ͉ amino acid discrimination ͉ editing ͉ genetic code accuracy ͉ ATP binding A minoacyl-tRNA synthetases (AARSs) establish the genetic code by attaching specific amino acids to the 3Ј ends of tRNAs bearing the anticodons cognate for those amino acids (1-3). These modular enzymes fall into two classes (I and II), based on the architectures of their catalytic domains (4). For both classes, aminoacylation proceeds in two steps catalyzed by one active site. A bound amino acid such as Ala is activated with ATP to form the enzyme-bound aminoacyl adenylate intermediate (Ala-AMP), releasing inorganic pyrophosphate (step 1). The aminoacyl moiety of the adenylate is then transferred to the 2Ј-or 3Ј-OH of the ribose of A76, the universally conserved 3Ј nucleotide of tRNA, with the release of AMP (step 2). Misactivation of amino acids occurs occasionally when a wrong amino acid fits into the active site, usually because it is smaller than the cognate one. Here, we describe structural features in a specific AARS (complexed with three different amino acids) that explain a longstanding paradox about activation of a larger amino acid.AlaRS ϩ Ala ϩ ATP ¡ AlaRS(Ala-AMP) ϩ PPi

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Section: Resultssupporting

confidence: 88%

Breaking sieve for steric exclusion of a noncognate amino acid from active site of a tRNA synthetase

Swairjo

Schimmel²

2005

Proc. Natl. Acad. Sci. U.S.A.

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“…In similar adenylation reactions it has been suggested that a general mechanism for substrate adenylation may involve conserved arginine or lysine residues and two or three divalent cations that stabilize the pyrophosphate leaving group and the pentavalent transition-state intermediate formed during adenylation (38,39). Although we have been unable to locate magnesium ions in the current EAS structures, such stabilizing ions have been identified in the other N-type ATP pyrophosphatases and are reported to be required for maximal AS activity (30).…”

Section: Resultsmentioning

confidence: 64%

Substrate Induced Conformational Changes in Argininosuccinate Synthetase

Lemke

Howell

2002

Journal of Biological Chemistry

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Argininosuccinate synthetase (AS) is the rate-limiting enzyme of both the urea and arginine-citrulline cycles. In mammals, deficiency of AS leads to citrullinemia, a debilitating and often fatal autosomal recessive urea cycle disorder, whereas its overexpression for sustained nitric oxide production via the arginine-citrulline cycle leads to the potentially fatal hypotension associated with septic and cytokine-induced circulatory shock. The crystal structures of Escherichia coli argininosuccinate synthetase (EAS) in complex with ATP and with ATP and citrulline have been determined at 2.0-Å resolution. These are the first EAS structures to be solved in the presence of a nucleotide substrate and clearly identify the residues that interact with both ATP and citrulline. Two distinct conformations are revealed for ATP, both of which are believed to be catalytically relevant. In addition, comparisons of these EAS structures with those of the apoenzyme and EAS complexed with aspartate and citrulline (Lemke, C. T., and Howell, P. L. (2001) Structure (Lond.) 9, 1153-1164) provide structural evidence of ATP-induced conformational changes in the nucleotide binding domain. Combined, these structures also provide structural explanations of some of the observed kinetic properties of the enzyme and have enabled a detailed enzymatic mechanism of AS catalysis to be proposed.Argininosuccinate synthetase (AS 1 ; EC 6.3.4.5) catalyzes the reversible conversion of citrulline, aspartate, and ATP to argininosuccinate, AMP, and inorganic pyrophosphate (Fig. 1). There are three important metabolic processes that require this catalysis. First, in all organisms AS catalyzes the penultimate step in the biosynthesis of arginine, one of the 20 amino acid building blocks of life, and a precursor for the synthesis of several other biomolecules. Second, AS participates in the urea cycle, a five-enzyme cycle that employs four of the enzymes of arginine biosynthesis to detoxify ammonia through the production of urea. Ammonia detoxification is critical for the survival of higher organisms. In humans, failure to produce functional AS leads to the buildup of citrulline, ammonia, and orotic acid. If untreated, the neurotoxic ammonia can cause brain damage and coma, and in cases where urea cycle function is significantly compromised (less than 5% activity) the condition is typically fatal (1). Finally, AS and a second urea cycle enzyme, argininosuccinate lyase, together with the flavoprotein nitricoxide synthase form the arginine-citrulline cycle, an abbreviated urea cycle that provides de novo arginine biosynthesis for sustainable overproduction of nitric oxide (NO (2, 3)). NO is a small, membrane-permeable, highly reactive molecule that plays key roles in a wide range of mammalian processes including blood pressure control, neurotransmission, apoptosis, immune system function, and wound healing (for reviews, see . AS is the rate-limiting enzyme in both the urea and the arginine-citrulline cycles (11,12) and is therefore a key participant in all thes...

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“…The 3Ј-OH of the ribose forms a hydrogen bond with the backbone carbonyl of Thr281. The triphosphate bends over the adenosine moiety, a conformation observed in the yeast AspRS (Cavarelli et al, 1994) and SerRS from T. thermophilus (Belrhali et al, 1995) and may be unique to class II aaRS ) bridge the ␤-and ␥-phosphates , which has also been observed with other class II aaRS such as SerRS and AspRS (Schmitt et al, 1998). Two Arg residues, one contributed by the motif 2 loop and the other by the motif 3 helix, interact with the ␥-phosphate.…”

Section: Crystal Structure Of Histidyl-trna Synthetasementioning

confidence: 76%

Histidyl-tRNA Synthetase

Freist¹,

Jf²,

Rühlmann³

et al. 1999

Biological Chemistry

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Histidyl-tRNA synthetase (HisRS) is responsible for the synthesis of histidyl-transfer RNA, which is essential for the incorporation of histidine into proteins. This amino acid has uniquely moderate basic properties and is an important group in many catalytic functions of enzymes.A compilation of currently known primary structures of HisRS shows that the subunits of these homodimeric enzymes consist of 420 -550 amino acid residues. This represents a relatively short chain length among aminoacyl-tRNA synthetases (aaRS), whose peptide chain sizes range from about 300 to 1100 amino acid residues.The crystal structures of HisRS from two organisms and their complexes with histidine, histidyl-adenylate and histidinol with ATP have been solved. HisRS from Escherichia coli and Thermus thermophilus are very similar dimeric enzymes consisting of three domains: the N-terminal catalytic domain containing the sixstranded antiparallel ␤-sheet and the three motifs characteristic of class II aaRS, a HisRS-specific helical domain inserted between motifs 2 and 3 that may contact the acceptor stem of the tRNA, and a C-terminal ␣/␤ domain that may be involved in the recognition of the anticodon stem and loop of tRNA His .The aminoacylation reaction follows the standard two-step mechanism. HisRS also belongs to the group of aaRS that can rapidly synthesize diadenosine tetraphosphate, a compound that is suspected to be involved in several regulatory mechanisms of cell metabolism. Many analogs of histidine have been tested for their properties as substrates or inhibitors of HisRS, leading to the elucidation of structure-activity relationships concerning configuration, importance of the carboxy and amino group, and the nature of the side chain.HisRS has been found to act as a particularly important antigen in autoimmune diseases such as rheumatic arthritis or myositis. Successful attempts have been made to identify epitopes responsible for the complexation with such auto-antibodies.

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The structural basis for seryl-adenylate and Ap4A synthesis by seryl-tRNA synthetase

Cited by 87 publications

References 40 publications

Breaking sieve for steric exclusion of a noncognate amino acid from active site of a tRNA synthetase

Breaking sieve for steric exclusion of a noncognate amino acid from active site of a tRNA synthetase

Substrate Induced Conformational Changes in Argininosuccinate Synthetase

Histidyl-tRNA Synthetase

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