Chemically modified ATP derivatives for the study of aminoacyl-tRNA synthetases from baker's yeast: ATP analogs with fixed conformations or modified triphosphate chains in the aminoacylation reaction

Freist, Wolfgang; Wiedner, Harald; Cramer, Friedrich

doi:10.1016/0045-2068(80)90009-7

Cited by 14 publications

(5 citation statements)

References 37 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Depending upon the particular synthetase, /3,7-methylene-and imido-ATP analogues either are substrates or inhibitors or have little effect (Papas & Case, 1970;Freist et al, 1980). Like-wise, El enzymes from diverse sources may differ with respect to their nucleotide specificities, although the highly conserved nature of the ubiquitin system [see Jentsch et al (1990) and references cited therein] suggests otherwise.…”

Section: Discussionmentioning

confidence: 99%

Uncoupling ubiquitin-protein conjugation from ubiquitin-dependent proteolysis by use of .beta.,.gamma.-nonhydrolyzable ATP analogs

Johnston

Cohen²

1991

Biochemistry

View full text Add to dashboard Cite

Pathways of ubiquitin-dependent protein degradation have in common two requirements for ATP. Ubiquitin activation by the enzyme E1 is accompanied by ATP hydrolysis to yield AMP and PPi, and during conjugate breakdown, the ubiquitin-dependent protease hydrolyzes ATP to ADP and Pi. We show here that either of two beta, gamma-nonhydrolyzable ATP analogues, 5'-adenylyl imidodiphosphate or 5'-adenylyl methylenediphosphate, can support ubiquitin-protein conjugation. With the ubiquitin-dependent protease, however, neither analogue could substitute for ATP. Thus, the substitution of a beta, gamma-nonhydrolyzable analogue for ATP offers a simple method to uncouple ubiquitin conjugation from proteolysis in crude systems. On the basis of pyrophosphate exchange kinetics, E1 has apparent Km and Vmax values that are similar for ATP and the analogues, but substrate inhibition by 5'-adenylyl methylenediphosphate made use of the beta, gamma-imido analogue preferable. In one application, beta, gamma-imido-ATP was used in combination with ubiquitin aldehyde (an inhibitor of ubiquitin-protein isopeptidases) to establish that several unfolded RNase A derivatives are recognized equally as ubiquitination substrates. This result extends an earlier study [Dunten, R. L., & Cohen, R. E. (1989) J. Biol. Chem. 264, 16739-16747] to show that conjugate yields, upon which relative ubiquitination rates were based, were not influenced by differential ubiquitin-dependent proteolysis. In a second application, ATP and beta, gamma-imido-ATP were compared in a pulse-chase experiment to investigate the contributions of ATP-dependent proteolysis and isopeptidase activities to conjugate stability.

show abstract

Section: Discussionmentioning

confidence: 99%

Uncoupling ubiquitin-protein conjugation from ubiquitin-dependent proteolysis by use of .beta.,.gamma.-nonhydrolyzable ATP analogs

Johnston

Cohen²

1991

Biochemistry

View full text Add to dashboard Cite

show abstract

“…Following initial unsuccessful attempts to generate crystals in the presence of ATP and Asn, CD Hs AsnRS was incubated in the presence of Asn and the ATP analog, AMPPNP 25 . Diffraction quality crystals were obtained; and we solved the structure (refined at 2.2 Å resolution), revealing the presence of Asn-AMP in the active site, presumably formed by attack of the amino acid on the α-phosphate (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Initial attempts to crystallize CD Hs AsnRS in the presence of ATP and Asn, with a view to capturing the complex with the activated Asn-AMP intermediate were not successful. Therefore, we employed the ATP analog, AMPPNP, which can serve as a substrate for some aminoacyl tRNA synthetases 25 , 44 . Diffraction quality crystals were obtained under these conditions.…”

Section: Discussionmentioning

confidence: 99%

Reaction hijacking inhibition of Plasmodium falciparum asparagine tRNA synthetase

Xie,

Wang,

Morton

et al. 2024

Nat Commun

View full text Add to dashboard Cite

Malaria poses an enormous threat to human health. With ever increasing resistance to currently deployed drugs, breakthrough compounds with novel mechanisms of action are urgently needed. Here, we explore pyrimidine-based sulfonamides as a new low molecular weight inhibitor class with drug-like physical parameters and a synthetically accessible scaffold. We show that the exemplar, OSM-S-106, has potent activity against parasite cultures, low mammalian cell toxicity and low propensity for resistance development. In vitro evolution of resistance using a slow ramp-up approach pointed to the Plasmodium falciparum cytoplasmic asparaginyl-tRNA synthetase (PfAsnRS) as the target, consistent with our finding that OSM-S-106 inhibits protein translation and activates the amino acid starvation response. Targeted mass spectrometry confirms that OSM-S-106 is a pro-inhibitor and that inhibition of PfAsnRS occurs via enzyme-mediated production of an Asn-OSM-S-106 adduct. Human AsnRS is much less susceptible to this reaction hijacking mechanism. X-ray crystallographic studies of human AsnRS in complex with inhibitor adducts and docking of pro-inhibitors into a model of Asn-tRNA-bound PfAsnRS provide insights into the structure-activity relationship and the selectivity mechanism.

show abstract

“…23 1; bulky substituents are tolerated in Position 2 but not in Position 8. The generalization can be made that phenylalanyl-tRNA synthetases may accept ATP in the form of an ATP-Mg 2 ® complex in the anti conformation with a coordination of the cation to the N-7 as assumed for the E. coli and the yeast cytoplasmic phenylalanyl-tRNA synthetase [12,20,24]. Valyl-and isoleucyl-tRNA synthetases from yeast on the other hand perhaps may accept ATP in the syn conformation' 13 L It is also assumed, and is probably a common feature for phenylalanyltRNA synthetases that the pyrophosphate moiety within the Michaelis-Menten complex is orientated by this magnesium cation and that the amino acid can best approach the α-phosphorus from downsite of the sugar plane' 12 ' 25 1 Another common feature is the unimportance of the 2'-hydroxyl group for catalysis.…”

Section: Resultsmentioning

confidence: 99%

Phenylalanyl-tRNA Synthetases from Hen Liver Cytoplasm and Mitochondria, Yeast Cytoplasm and Mitochondria, and fromEscherichia coli: Substrate Specificity Relationship with regard to ATP Analogs

Gabius¹,

Freist²,

Cramer³

1982

Hoppe-Seyler´s Zeitschrift für physiologische Chemie

Self Cite

View full text Add to dashboard Cite

Twelve structural analogs of ATP have been tested in the aminoacylation reaction of phenylalanyl-tRNA synthetases from hen liver cytoplasm and mitochondria, yeast cytoplasm and mitochondria and E. coli. Three compounds are substrates for all five phenylalanyl-tRNA synthetases, three are completely inactive, while the other ATP analogs show differing properties with the different enzymes. Their K m , KI and V values have been determined.

show abstract

Chemically modified ATP derivatives for the study of aminoacyl-tRNA synthetases from baker's yeast: ATP analogs with fixed conformations or modified triphosphate chains in the aminoacylation reaction

Cited by 14 publications

References 37 publications

Uncoupling ubiquitin-protein conjugation from ubiquitin-dependent proteolysis by use of .beta.,.gamma.-nonhydrolyzable ATP analogs

Uncoupling ubiquitin-protein conjugation from ubiquitin-dependent proteolysis by use of .beta.,.gamma.-nonhydrolyzable ATP analogs

Reaction hijacking inhibition of Plasmodium falciparum asparagine tRNA synthetase

Phenylalanyl-tRNA Synthetases from Hen Liver Cytoplasm and Mitochondria, Yeast Cytoplasm and Mitochondria, and fromEscherichia coli: Substrate Specificity Relationship with regard to ATP Analogs

Contact Info

Product

Resources

About