Rational Design of Femtomolar Inhibitors of Isoleucyl tRNA Synthetase from a Binding Model for Pseudomonic Acid-A

Brown, Murray J. B.; Mensah, Lucy; Doyle, Michael L.; Broom, Nigel J. P.; Osbourne, Neal; Forrest, Andrew K.; Richardson, Christine M.; O’Hanlon, Peter J.; Pope, Andrew J.

doi:10.1021/bi000148v

Cited by 72 publications

(56 citation statements)

References 29 publications

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“…Given that N-acylsulfonamide derivatives are high-affinity, slow-binding inhibitors of isoleucyl-tRNA synthetases (158)(159)(160) and have "drug-like" physical properties (161,162), a sulfonamide analog of the βAspAMP intermediate (6) (Figure 6) was also prepared and characterized for its ability to inhibit recombinant, wild-type human ASNS (24). These in vitro experiments clearly showed that sulfonamide derivative 6 is a slow-onset, tight-binding inhibitor that interacts with the free enzyme, and revealed the importance of carboxyl and amino groups in the recognition and binding of this class of βAspAMP analog.…”

Section: Sulfonamide Derivatives As Inhibitors Of Human Asparagine Symentioning

confidence: 99%

Asparagine Synthetase Chemotherapy

Richards

Kilberg

2006

Annu. Rev. Biochem.

204

221

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Modern clinical treatments of childhood acute lymphoblastic leukemia (ALL) employ enzymebased methods for depletion of blood asparagine in combination with standard chemotherapeutic agents. Significant side effects can arise in these protocols and, in many cases, patients develop drug-resistant forms of the disease that may be correlated with up-regulation of the enzyme glutamine-dependent asparagine synthetase (ASNS). Though the precise molecular mechanisms that result in the appearance of drug resistance are the subject of active study, potent ASNS inhibitors may have clinical utility in treating asparaginase-resistant forms of childhood ALL. This review provides an overview of recent developments in our understanding of (a) the structure and catalytic mechanism of ASNS, and (b) the role that ASNS may play in the onset of drug-resistant childhood ALL. In addition, the first successful, mechanism-based efforts to prepare and characterize nanomolar ASNS inhibitors are discussed, together with the implications of these studies for future efforts to develop useful drugs.

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Section: Sulfonamide Derivatives As Inhibitors Of Human Asparagine Symentioning

confidence: 99%

Asparagine Synthetase Chemotherapy

Richards

Kilberg

2006

Annu. Rev. Biochem.

204

221

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“…Until the discovery of a class I lysyl-tRNA synthetase (LysRS1) (4), it was believed that each amino acid was exclusively served by a single aaRS of either class I or class II. Although the aaRSs are of interest from many perspectives, including their mechanism of catalysis and proofreading (1), the intricacies of protein-RNA interactions (5), their involvement in cellular signaling functions (6), their practical applicability as potential antimicrobial targets (7), their probable role in genetic diseases (8,9), or the ability to engineer aaRSs to cotranslationally introduce unnatural amino acids into proteins (10), there remains the fundamental question about the role of these proteins in the evolution of the genetic code. The central dilemma is whether the evolution of the aaRSs is the evolution of the genetic code itself, or whether the tRNA synthetases emerged in the context of a previously established code.…”

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confidence: 99%

Emergence of the universal genetic code imprinted in an RNA record

Hohn¹,

Park²,

O’Donoghue³

et al. 2006

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

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The molecular basis of the genetic code manifests itself in the interaction of the aminoacyl-tRNA synthetases and their cognate tRNAs. The fundamental biological question regarding these enzymes' role in the evolution of the genetic code remains open. Here we probe this question in a system in which the same tRNA species is aminoacylated by two unrelated synthetases. Should this tRNA possess major identity elements common to both enzymes, this would favor a scenario where the aminoacyl-tRNA synthetases evolved in the context of preestablished tRNA identity, i.e., after the universal genetic code emerged. An experimental system is provided by the recently discovered O-phosphoseryl-tRNA synthetase (SepRS), which acylates tRNA Cys with phosphoserine (Sep), and the well known cysteinyl-tRNA synthetase, which charges the same tRNA with cysteine. We determined the identity elements of Methanocaldococcus jannaschii tRNA Cys in the aminoacylation reaction for the two Methanococcus maripaludis synthetases SepRS (forming Sep-tRNA Cys ) and cysteinyl-tRNA synthetase (forming Cys-tRNA Cys ). The major elements, the discriminator base and the three anticodon bases, are shared by both tRNA synthetases. An evolutionary analysis of archaeal, bacterial, and eukaryotic tRNA Cys sequences predicted additional SepRS-specific minor identity elements (G37, A47, and A59) and suggested the dominance of vertical inheritance for tRNA Cys from a single common ancestor. Transplantation of the identified identity elements into the Escherichia coli tRNA Gly scaffold endowed facile phosphoserylation activity on the resulting chimera. Thus, tRNA Cys identity is an ancient RNA record that depicts the emergence of the universal genetic code before the evolution of the modern aminoacylation systems.aminoacyl-tRNA synthetase ͉ evolution ͉ O-phosphoseryl-tRNA synthetase ͉ tRNA identity

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“…Aminoacyl-tRNA synthetases have gained attention recently as potential targets for antibiotics (1)(2)(3)(4)(5)(6)(7)(8)(9)(10). Identifying differences in the catalytic mechanisms of bacterial and human aminoacyl-tRNA synthetases will facilitate the development of antibiotics that selectively target the bacterial aminoacyltRNA synthetases.…”

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confidence: 99%

“…Tyrosyl-tRNA synthetase catalyzes the attachment of tyrosine to tyrosine tRNA (tRNA Tyr ) 1 by an ATP-dependent twostep reaction mechanism. In the first step, tyrosine is activated by MgATP to form an enzyme-bound tyrosyl-adenylate intermediate.…”

mentioning

confidence: 99%

Potassium Functionally Replaces the Second Lysine of the KMSKS Signature Sequence in Human Tyrosyl-tRNA Synthetase

Austin¹,

First

2002

Journal of Biological Chemistry

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Unlike their bacterial homologues, a number of eukaryotic tyrosyl-tRNA synthetases require potassium to catalyze the aminoacylation reaction. In addition, the second lysine in the class I-specific KMSKS signature motif is absent from all known eukaryotic tyrosyl-tRNA synthetase sequences, except those of higher plants. This lysine, which is the most highly conserved residue in the class I aminoacyl-tRNA synthetase family, has been shown to interact with the pyrophosphate moiety of the ATP substrate in the Bacillus stearothermophilus tyrosyl-tRNA synthetase. Equilibrium dialysis and presteady-state kinetic analyses were used to determine the role that potassium plays in the tyrosine activation reaction in the human tyrosyl-tRNA synthetase and whether it can be replaced by any of the other alkali metals. Kinetic analyses indicate that potassium interacts with the pyrophosphate moiety of ATP, stabilizing the E⅐Tyr⅐ATP and E⅐[Tyr-ATP] ‡ complexes by 2.3 and 4.3 kcal/mol, respectively. Potassium also appears to stabilize the asymmetric conformation of the human tyrosyltRNA synthetase dimer by 0.7 kcal/mol. Rubidium is the only other alkali metal that can replace potassium in catalyzing tyrosine activation, although the forward rate constant is half of that observed when potassium is present. The above results are consistent with the hypothesis that potassium functionally replaces the second lysine in the KMSKS signature sequence. Possible implications of these results with respect to the design of antibiotics that target bacterial aminoacyl-tRNA synthetases are discussed.Aminoacyl-tRNA synthetases have gained attention recently as potential targets for antibiotics (1-10). Identifying differences in the catalytic mechanisms of bacterial and human aminoacyl-tRNA synthetases will facilitate the development of antibiotics that selectively target the bacterial aminoacyltRNA synthetases. We are currently investigating the catalytic mechanisms of the human and Bacillus stearothermophilus tyrosyl-tRNA synthetases to elucidate the differences between these two enzymes.Tyrosyl-tRNA synthetase catalyzes the attachment of tyrosine to tyrosine tRNA (tRNA Tyr ) 1 by an ATP-dependent twostep reaction mechanism. In the first step, tyrosine is activated by MgATP to form an enzyme-bound tyrosyl-adenylate intermediate. The second step consists of the transfer of tyrosine to the 3Ј end of tRNA Tyr .TyrRS ϩ Tyr ϩ MgATP^TyrRS ⅐ Tyr-AMP ϩ PP i (Eq. 1)Tyrosyl-tRNA synthetase is a homodimer that displays "halfof-the-sites" reactivity, with only one tyrosyl-adenylate molecule formed per dimer (11-13). Although most of the active site amino acids are conserved between the human and B. stearothermophilus tyrosyl-tRNA synthetases, several differences exist between the two enzymes, including the inability of the human and bacterial tyrosyl-tRNA synthetases to aminoacylate each other's tRNA Tyr (14,15). In addition, sequence analyses indicate that the human tyrosyl-tRNA synthetase is Ͻ16% identical to the B. stearothermophilus enzyme and that four...

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Rational Design of Femtomolar Inhibitors of Isoleucyl tRNA Synthetase from a Binding Model for Pseudomonic Acid-A

Cited by 72 publications

References 29 publications

Asparagine Synthetase Chemotherapy

Asparagine Synthetase Chemotherapy

Emergence of the universal genetic code imprinted in an RNA record

Potassium Functionally Replaces the Second Lysine of the KMSKS Signature Sequence in Human Tyrosyl-tRNA Synthetase

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