Substrate-assisted catalysis of peptide bond formation by the ribosome

Weinger, Joshua S.; Parnell, K. Mark; Dorner, Silke; Green, Rachel; Strobel, Scott A.

doi:10.1038/nsmb841

Cited by 271 publications

(346 citation statements)

References 39 publications

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“…Guided by PTC components, the rotatory motion facilitates peptide-bond formation and nascent-chain elongation. Furthermore, this motion places the A-site nucleophilic amine and the P-site carbonyl carbon at a distance allowing for interactions with the P-site tRNA A76 O2Ј throughout a significant part of the rotatory motion (3), consistent with its suggested participation in peptide-bond formation catalysis (8). The nascent proteins are directed by the rotatory motion into the exit tunnel at extended conformation, fitting the tunnel's narrow opening.…”

mentioning

confidence: 66%

The transition state for formation of the peptide bond in the ribosome

Gindulytė

Bashan

Agmon

et al. 2006

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

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Using quantum mechanics and exploiting known crystallographic coordinates of tRNA substrate located in the ribosome peptidyl transferase center around the 2-fold axis, we have investigated the mechanism for peptide-bond formation. The calculation is based on a choice of 50 atoms assumed to be important in the mechanism. We used density functional theory to optimize the geometry and energy of the transition state (TS) for peptide-bond formation. The TS is formed simultaneously with the rotatory motion enabling the translocation of the A-site tRNA 3′ end into the P site, and we estimated the magnitude of rotation angle between the A-site starting position and the place at which the TS occurs. The calculated TS activation energy, E a , is 35.5 kcal (1 kcal = 4.18 kJ)/mol, and the increase in hydrogen bonding between the rotating A-site tRNA and ribosome nucleotides as the TS forms appears to stabilize it to a value qualitatively estimated to be ≈18 kcal/mol. The optimized geometry corresponds to a structure in which the peptide bond is being formed as other bonds are being broken, in such a manner as to release the P-site tRNA so that it may exit as a free molecule and be replaced by the translocating A-site tRNA. At TS formation the 2′ OH group of the P-site tRNA A76 forms a hydrogen bond with the oxygen atom of the carboxyl group of the amino acid attached to the A-site tRNA, which may be indicative of its catalytic role, consistent with recent biochemical experiments.

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mentioning

confidence: 66%

The transition state for formation of the peptide bond in the ribosome

Gindulytė

Bashan

Agmon

et al. 2006

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

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“…It seems beyond doubt that the proton transfer in this reaction has to occur via the O2′ of the P-site substrate A76 ribose (22). The easiest way in which this could happen is through a six-membered transition state where the O2′ acts as a proton shuttle (15,16,20,28).…”

Section: Discussionmentioning

confidence: 99%

“…Recent years have witnessed considerable progress in our understanding of the peptidyl transfer process due to high-resolution crystallographic structures of the large ribosomal subunit with transition state (TS) analogs (1), as well as kinetic measurements (2)(3)(4)(5)(6)(7)(8)(9), mutagenesis data (4,(10)(11)(12)(13)(14), and computational studies (15)(16)(17). Hence, the current model of the peptidyl transfer reaction is that ribosomal nucleobases are not directly involved in bond making or breaking through acid-base catalysis (4,10,11,13,18,19), but that the A76 2′-OH group of the P-site substrate plays a key role in mediating proton transfer from the attacking nucleophile to the leaving 3′ ester oxygen (1,15,(20)(21)(22). Furthermore, it was shown that the lower free energy barrier for the ribosome reaction, compared to an uncatalyzed reference reaction in water, is entirely due to a less negative activation entropy (3,6,7).…”

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confidence: 99%

The transition state for peptide bond formation reveals the ribosome as a water trap

Wallin

Åqvist

2010

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

103

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Recent progress in elucidating the peptide bond formation process on the ribosome has led to notion of a proton shuttle mechanism where the 2'-hydroxyl group of the P-site tRNA plays a key role in mediating proton transfer between the nucleophile and leaving group, whereas ribosomal groups do not actively participate in the reaction. Despite these advances, the detailed nature of the transition state for peptidyl transfer and the role of several trapped water molecules in the peptidyl transferase center remain major open questions. Here, we employ high-level quantum chemical ab initio calculations to locate and characterize global transition states for the reaction, described by a molecular model encompassing all the key elements of the reaction center. The calculated activation enthalpy as well as structures are in excellent agreement with experimental data and point to feasibility of an eight-membered "double proton shuttle" mechanism in which an auxiliary water molecule, observed both in computer simulations and crystal structures, actively participates. A second conserved water molecule is found to be of key importance for stabilizing developing negative charge on the substrate oxyanion and its presence is catalytically favorable both in terms of activation enthalpy and entropy. Transition states calculated both for six-and eight-membered mechanisms are invariably late and do not involve significant charge development on the attacking amino group. Predicted kinetic isotope effects consistent with this picture are similar to those observed for uncatalyzed ester aminolysis reactions in solution.density functional theory | protein synthesis T he peptide bond formation step in protein synthesis is catalyzed by the peptidyl transferase center (PTC) on the large ribosomal subunit. This reaction involves the attack of the α-amino group of an aminoacyl-tRNA molecule bound to ribosomal A-site on the ester carbon of the peptidyl-tRNA in the adjacent P-site of the ribosome. The growing peptide chain is thereby transferred to the A-site tRNA and elongated by one amino acid. Recent years have witnessed considerable progress in our understanding of the peptidyl transfer process due to high-resolution crystallographic structures of the large ribosomal subunit with transition state (TS) analogs (1), as well as kinetic measurements (2-9), mutagenesis data (4, 10-14), and computational studies (15)(16)(17). Hence, the current model of the peptidyl transfer reaction is that ribosomal nucleobases are not directly involved in bond making or breaking through acid-base catalysis (4,10,11,13,18,19), but that the A76 2′-OH group of the P-site substrate plays a key role in mediating proton transfer from the attacking nucleophile to the leaving 3′ ester oxygen (1,15,(20)(21)(22). Furthermore, it was shown that the lower free energy barrier for the ribosome reaction, compared to an uncatalyzed reference reaction in water, is entirely due to a less negative activation entropy (3, 6, 7). Besides contributions from substrate proximity and...

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“…Ribozymes are catalytic RNA molecules, some of which catalyse extremely important cellular reactions, notably processing tRNA and mRNA (1) and condense amino acids to form polypeptides (2)(3)(4)(5). Ribozymes are very widespread; for example ribonuclease P (6, 7) processes precursor tRNA in all domains of life.…”

mentioning

confidence: 99%