A substrate binding model for the KEOPS tRNA modifying complex

Beenstock, Jonah; Ona, Samara Mishelle; Porat, Jennifer; Orlicky, Stephen; Wan, Leo C. K.; Ceccarelli, Derek F.; Maisonneuve, Pierre; Szilard, Rachel K.; Yin, Zhe; Setiaputra, Dheva; Mao, Daniel Y.L.; Khan, Morgan F.; Raval, Shaunak; Schriemer, David C.; Bayfield, Mark A.; Durocher, Daniel; Sicheri, Frank

doi:10.1038/s41467-020-19990-5

Cited by 27 publications

(91 citation statements)

References 68 publications

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“…In line with this prediction, the chief function of Bud32 appears directed at ATP hydrolysis (i.e. transfer of phosphate to water) ( 102 ).…”

Section: How Keops Functions To Modify Trnamentioning

confidence: 65%

“…The binding mode between Bud32 and Cgi121 is reminiscent of cyclin-dependent kinases binding to cyclins ( 112 ). Thus, not unexpectedly, Cgi121 potentiates both the ATPase and low level autophosphorylation activity of Bud32 (note that the autophosphorylation activity is unlikely to be physiologically relevant) ( 9 , 101 , 102 ). Structures of Bud32 in complex with Cgi121 reveal an active-like conformation with productively positioned catalytic elements ( 99 , 101 ).…”

Section: How Keops Functions To Modify Trnamentioning

confidence: 93%

“…Accordingly, Bud32 proteins manifest autophosphorylation activity ( 5 , 8 , 9 ) and its phospho-transfer function is essential for biological function ( 9 , 11 ). However, the absence of structural elements normally required for protein phosphorylation as well as experiments showing that the autophosphorylation acceptor-sites are not required for physiological activity ( 7 , 102 ) suggests an alternate role for its phospho-transfer function. In line with this prediction, the chief function of Bud32 appears directed at ATP hydrolysis (i.e.…”

Section: How Keops Functions To Modify Trnamentioning

confidence: 99%

“…The highly conserved C-terminal tail of Bud32 interacts with the active site of Kae1 (Figure 4A ) ( 8 , 9 ). While not essential for binding to Kae1, the C-terminal tail of Bud32 is required the ability of KEOPS to modify tRNA in vitro and for KEOPS functionality in vivo as assessed in yeast ( 9 , 102 ).…”

Section: How Keops Functions To Modify Trnamentioning

confidence: 99%

“…This ability is essential for the efficient recruitment of tRNA to KEOPS. A co-crystal structure of a Cgi121-tRNA complex revealed that tRNA binding by Cgi121 is directed almost exclusively towards the 3' CCA tail (positions 74–76, 5'-C 74 -C 75 -A 76 -3') (Figure 4B ) ( 102 ). The binding surface on Cgi121 for tRNA is composed primarily of α−helices 1, 3 and 4.…”

Section: How Keops Functions To Modify Trnamentioning

confidence: 99%

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The structural and functional workings of KEOPS

Beenstock

Sicheri

2021

Nucleic Acids Research

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KEOPS (Kinase, Endopeptidase and Other Proteins of Small size) is a five-subunit protein complex that is highly conserved in eukaryotes and archaea and is essential for the fitness of cells and for animal development. In humans, mutations in KEOPS genes underlie Galloway–Mowat syndrome, which manifests in severe microcephaly and renal dysfunction that lead to childhood death. The Kae1 subunit of KEOPS catalyzes the universal and essential tRNA modification N6-threonylcarbamoyl adenosine (t6A), while the auxiliary subunits Cgi121, the kinase/ATPase Bud32, Pcc1 and Gon7 play a supporting role. Kae1 orthologs are also present in bacteria and mitochondria but function in distinct complexes with proteins that are not related in structure or function to the auxiliary subunits of KEOPS. Over the past 15 years since its discovery, extensive study in the KEOPS field has provided many answers towards understanding the roles that KEOPS plays in cells and in human disease and how KEOPS carries out these functions. In this review, we provide an overview into recent advances in the study of KEOPS and illuminate exciting future directions.

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“…In line with this prediction, the chief function of Bud32 appears directed at ATP hydrolysis (i.e. transfer of phosphate to water) ( 102 ).…”

Section: How Keops Functions To Modify Trnamentioning

confidence: 65%

Section: How Keops Functions To Modify Trnamentioning

confidence: 93%

Section: How Keops Functions To Modify Trnamentioning

confidence: 99%

Section: How Keops Functions To Modify Trnamentioning

confidence: 99%

Section: How Keops Functions To Modify Trnamentioning

confidence: 99%

See 3 more Smart Citations

The structural and functional workings of KEOPS

Beenstock

Sicheri

2021

Nucleic Acids Research

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The archaeal KEOPS complex possesses a functional Gon7 homolog and has an essential function independent of the cellular t⁶A modification level

Gan

Zhang

et al. 2023

mLife

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Kinase, putative Endopeptidase, and Other Proteins of Small size (KEOPS) is a multisubunit protein complex conserved in eukaryotes and archaea. It is composed of Pcc1, Kae1, Bud32, Cgi121, and Gon7 in eukaryotes and is primarily involved in N 6 -threonylcarbamoyl adenosine (t 6 A) modification of transfer RNAs (tRNAs). Recently, it was reported that KEOPS participates in homologous recombination (HR) repair in yeast. To characterize the KEOPS in archaea (aKEOPS), we conducted genetic and biochemical analyses of its encoding genes in the hyperthermophilic archaeon Saccharolobus islandicus. We show that aKEOPS also possesses five subunits, Pcc1, Kae1, Bud32, Cgi121, and Pcc1-like (or Gon7-like), just like eukaryotic KEOPS. Pcc1-like has physical interactions with Kae1 and Pcc1 and can mediate the monomerization of the dimeric subcomplex (Kae1-Pcc1-Pcc1-Kae1), suggesting that Pcc1-like is a functional homolog of the eukaryotic Gon7 subunit. Strikingly, none of the genes encoding aKEOPS subunits, including Pcc1 and Pcc1-like, can be deleted in the wild type and in a t 6 A modification complementary strain named TsaKI, implying that the aKEOPS complex is essential for an additional cellular process in this archaeon. Knock-down of the Cgi121 subunit leads to severe growth retardance in the wild type that is partially rescued in TsaKI. These results suggest that aKEOPS plays an essential role independent of the cellular t 6 A modification level. In addition, archaeal Cgi121 possesses dsDNA-binding activity that relies on its tRNA 3ʹ CCA tail binding module. Our study clarifies the subunit organization of archaeal KEOPS and suggests an origin of eukaryotic Gon7. The study also reveals a possible link between the function in t 6 A modification and the additional function, presumably HR.

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A suite of in vitro and in vivo assays for monitoring the activity of the pseudokinase Bud32

Beenstock

Ona

Sicheri

2022

Methods in Enzymology

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A substrate binding model for the KEOPS tRNA modifying complex

Cited by 27 publications

References 68 publications

The structural and functional workings of KEOPS

The structural and functional workings of KEOPS

The archaeal KEOPS complex possesses a functional Gon7 homolog and has an essential function independent of the cellular t⁶A modification level

A suite of in vitro and in vivo assays for monitoring the activity of the pseudokinase Bud32

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A substrate binding model for the KEOPS tRNA modifying complex

Cited by 27 publications

References 68 publications

The structural and functional workings of KEOPS

The structural and functional workings of KEOPS

The archaeal KEOPS complex possesses a functional Gon7 homolog and has an essential function independent of the cellular t6A modification level

A suite of in vitro and in vivo assays for monitoring the activity of the pseudokinase Bud32

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Product

Resources

About

The archaeal KEOPS complex possesses a functional Gon7 homolog and has an essential function independent of the cellular t⁶A modification level