Crystal Structure of the Apo Forms of ψ 55 tRNA Pseudouridine Synthase from Mycobacterium tuberculosis

Chaudhuri, Barnali N.; Chan, Sammy; Perry, Linda L.; Yeates, Todd O.

doi:10.1074/jbc.m401045200

Cited by 24 publications

(28 citation statements)

References 43 publications

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“…The shorter thumb loop in Cbf5 largely is disordered in the H/ACA RNP (30) (Fig. 5C), as is the case for apoTruB (35,42) as well as Cbf5-Nop10 (27-29). It has been proposed that this loop would become ordered when substrate binds, interacting with the major groove of PS1 (30), as illustrated in Fig.…”

Section: Discussionmentioning

confidence: 99%

H/ACA small nucleolar RNA pseudouridylation pockets bind substrate RNA to form three-way junctions that position the target U for modification

Feigon

2007

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

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), the authors note that in Fig. 1A, the sequence of the 5Ј side of the 5Ј pseudouridylation pocket of the U65 H/ACA snoRNA, as well as a portion of the lower stem of the 3Ј hairpin, were drawn incorrectly. Additionally, in the Fig. 1 legend and elsewhere in the text, the numbering of the substrate RNA, which was based on Bortolin et al. [Bortolin M-L, Ganot P, Kiss T (1999) EMBO J 18:457-469], differs by one nucleotide from that found in the genomic sequence (refer to the snoRNA database, snoRNA-LMBE-db, at www-snorna.biotoul.fr). The correct numbers for the pseudouridylation sites are 4427 and 4373 for the 5Ј and 3Ј pseudouridylation pockets, respectively. The corrected figure and legend appear below. These errors do not affect the conclusions of the article. (U65hp), the substrate rRNA (S14), and the U65hp/S14 complex. S14 contains the sequence of human 28S rRNA (4422-4434), including U4427 (underlined), which is targeted for pseudouridylation, and an extra U (lowercase) at the 5Ј end.

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Section: Discussionmentioning

confidence: 99%

H/ACA small nucleolar RNA pseudouridylation pockets bind substrate RNA to form three-way junctions that position the target U for modification

Feigon

2007

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

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“…15,16 Crystal structures of representative members of each family have been determined and reveal that even though sequence conservation between any two sub-families is low, all Ψ synthases have a structurally conserved catalytic core. [16][17][18][19][20][21][22][23][24][25][26][27][28][29][30] The only structures of Ψ synthasesubstrate complexes published to date are those of TruB and RluA co-crystallized with small RNA stem-loops that are minimal substrates of the enzymes. 21,22,26,27 These structures show that the substrate RNA binds to a positively charged surface on the protein with the target base flipped out of the stem-loop and into an active site cleft next to an invariant catalytic Asp.…”

Section: Introductionmentioning

confidence: 99%

The Crystal Structure of E. coli rRNA Pseudouridine Synthase RluE

Pan

Stroud

et al. 2007

Journal of Molecular Biology

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Pseudouridine synthase RluE modifies U2457 in a stem of 23 S RNA in Escherichia coli. This modification is located in the peptidyl transferase center of the ribosome. We determined the crystal structures of the C-terminal, catalytic domain of E. coli RluE at 1.2 A resolution and of full-length RluE at 1.6 A resolution. The crystals of the full-length enzyme contain two molecules in the asymmetric unit and in both molecules the N-terminal domain is disordered. The protein has an active site cleft, conserved in all other pseudouridine synthases, that contains invariant Asp and Tyr residues implicated in catalysis. An electropositive surface patch that covers the active site cleft is just wide enough to accommodate an RNA stem. The RNA substrate stem can be docked to this surface such that the catalytic Asp is adjacent to the target base, and a conserved Arg is positioned to help flip the target base out of the stem into the enzyme active site. A flexible RluE specific loop lies close to the conserved region of the stem in the model, and may contribute to substrate specificity. The stem alone is not a good RluE substrate, suggesting RluE makes additional interactions with other regions in the ribosome.

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“…In terms of Y173, it has been suggested that, at least in TruB, the corresponding tyrosine (Y67 in E. coli TruB) is involved in maintaining the catalytic aspartate in the charged state through the maintenance of a salt bridge (Hoang and Ferré-D'Amaré 2001;Chaudhuri et al 2004). Our results with mutations at this position suggest that a tyrosine at this position is indeed critical, but there was detectable activity with all the mutants except Y173S.…”

Section: Comparison Of Trua and Pus1pmentioning

confidence: 99%

Partial activity is seen with many substitutions of highly conserved active site residues in human Pseudouridine synthase 1

Sibert

Fischel‐Ghodsian²,

Patton³

2008

RNA

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Pseudouridine synthase 1 (Pus1p) is an enzyme that converts uridine to Pseudouridine (C) in tRNA and other RNAs in eukaryotes. The active site of Pus1p is composed of stretches of amino acids that are highly conserved and it is hypothesized that mutation of select residues would impair the enzyme's ability to catalyze the formation of C. However, most mutagenesis studies have been confined to substitution of the catalytic aspartate, which invariably results in an inactive enzyme in all C synthases tested. To determine the requirements for particular amino acids at certain absolutely conserved positions in Pus1p, three residues (R116, Y173, R267) that correspond to amino acids known to compose the active site of TruA, a bacterial C synthase that is homologous to Pus1p, were mutated in human Pus1p (hPus1p). The effects of those mutations were determined with three different in vitro assays of pseudouridylation and several tRNA substrates. Surprisingly, it was found that each of these components of the hPus1p active site could tolerate certain amino acid substitutions and in fact most mutants exhibited some activity. The most active mutants retained near wild-type activity at positions 27 or 28 in the substrate tRNA, but activity was greatly reduced or absent at other positions in tRNA readily modified by wild-type hPus1p.

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Crystal Structure of the Apo Forms of ψ 55 tRNA Pseudouridine Synthase from Mycobacterium tuberculosis

Cited by 24 publications

References 43 publications

H/ACA small nucleolar RNA pseudouridylation pockets bind substrate RNA to form three-way junctions that position the target U for modification

H/ACA small nucleolar RNA pseudouridylation pockets bind substrate RNA to form three-way junctions that position the target U for modification

The Crystal Structure of E. coli rRNA Pseudouridine Synthase RluE

Partial activity is seen with many substitutions of highly conserved active site residues in human Pseudouridine synthase 1

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