Structural and biochemical characterization of the cytosolic wheat cyclophilin TaCypA-1

Sekhon, Simranjeet Singh; Kaur, Harsimran; Dutta, Tanima; Singh, Kundan; Kumari, Sumita; Kang, Seung Kyu; Park, Sung Goo; Park, Byoung Chul; Jeong, Dae Gwin; Pareek, Ashwani; Woo, Eui-Jeon; Singh, Prabhjeet; Yoon, Tae-Myung

doi:10.1107/s0907444912051529

Cited by 17 publications

(32 citation statements)

References 42 publications

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“…3; Table I). The k cat /K m value for the wild-type CsCyp (5.6 s 21 mM 21 ) is consistent with values determined for other Cyp proteins (Dornan et al, 1999;Laxa et al, 2007;Sekhon et al, 2013). In addition, while the C40S and C168S mutants were similarly inhibited by CsA, the double mutant was more strongly inhibited by CsA relative to the wild-type protein ( Fig.…”

Section: On the Structural (Dis)similarities Between Classical And DIsupporting

confidence: 73%

“…Although 3D structures of divergent Cyps have been reported, the functional role of their unique structural elements (i.e. divergent loop, invariable Cys pair, and stabilizing Glu) remained unclear (Dornan et al, 1999;Taylor et al, 2001;Sekhon et al, 2013). Here, we show that, instead of coordinating a metal ion with His-54, as proposed previously (Dornan et al, 1999;Taylor et al, 2001), Cys-40 and Cys-168 form a disulfide bridge that blocks protein activity.…”

Section: Cscyp Binds Selectively To a Ctd Heptad Repeatmentioning

confidence: 55%

“…The CsCyp active site, composed of 13 residues that are also responsible for CsA binding (Fig. 1A), is identical to that of Cyp3, CypA, and TaCypA-1 (Pflügl et al, 1993;Ke et al, 1994;Dornan et al, 1999;Sekhon et al, 2013). Structure alignment of CsCyp with TaCypA-1, the only other plant divergent Cyp with known threedimensional (3D) structure, showed no major structural differences (root mean square deviation = 0.43 Å).…”

Section: On the Structural (Dis)similarities Between Classical And DImentioning

confidence: 95%

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A Redox 2-Cys Mechanism Regulates the Catalytic Activity of Divergent Cyclophilins

et al. 2013

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The citrus (Citrus sinensis) cyclophilin CsCyp is a target of the Xanthomonas citri transcription activator-like effector PthA, required to elicit cankers on citrus. CsCyp binds the citrus thioredoxin CsTdx and the carboxyl-terminal domain of RNA polymerase II and is a divergent cyclophilin that carries the additional loop KSGKPLH, invariable cysteine (Cys) residues Cys-40 and Cys-168, and the conserved glutamate (Glu) Glu-83. Despite the suggested roles in ATP and metal binding, the functions of these unique structural elements remain unknown. Here, we show that the conserved Cys residues form a disulfide bond that inactivates the enzyme, whereas Glu-83, which belongs to the catalytic loop and is also critical for enzyme activity, is anchored to the divergent loop to maintain the active site open. In addition, we demonstrate that Cys-40 and Cys-168 are required for the interaction with CsTdx and that CsCyp binds the citrus carboxyl-terminal domain of RNA polymerase II YSPSAP repeat. Our data support a model where formation of the Cys-40-Cys-168 disulfide bond induces a conformational change that disrupts the interaction of the divergent and catalytic loops, via Glu-83, causing the active site to close. This suggests a new type of allosteric regulation in divergent cyclophilins, involving disulfide bond formation and a loop-displacement mechanism.

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Section: On the Structural (Dis)similarities Between Classical And DIsupporting

confidence: 73%

Section: Cscyp Binds Selectively To a Ctd Heptad Repeatmentioning

confidence: 55%

Section: On the Structural (Dis)similarities Between Classical And DImentioning

confidence: 95%

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A Redox 2-Cys Mechanism Regulates the Catalytic Activity of Divergent Cyclophilins

et al. 2013

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“…Modelling of the 3D structure of LRT2 was performed as previously described 54 through the SWISS-Model server (http://swissmodel.expasy.org) using the crystal structure of wheat TaCypA-1 (PDB ID: 4E1Q) as a template 34 . Molecular graphics were presented using PyMOL software (http://www.pymol.org/).…”

Section: Methodsmentioning

confidence: 99%

“…6a) and mutations in this residue of rice LRT2/OsCYP2 and moss PpDGT cause severely impaired auxin signalling 17,30 . Homologous structure modelling based on the crystal structure of wheat (Triticum aestivum) TaCypA-1 (PDB code: 4E1Q) 34 , a cyclophilin sharing 87% identity with LRT2, revealed that Gly 72 is located at the surface of LRT2 immediately adjacent to a b-sheet ( Supplementary Fig. 6b).…”

Section: Lrt2 Catalyses Cis/trans Isomerization Of Osiaa11 Peptidementioning

confidence: 99%

Peptidyl-prolyl isomerization targets rice Aux/IAAs for proteasomal degradation during auxin signalling

et al. 2015

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In plants, auxin signalling is initiated by the auxin-promoted interaction between the auxin receptor TIR1, an E3 ubiquitin ligase, and the Aux/IAA transcriptional repressors, which are subsequently degraded by the proteasome. Gain-of-function mutations in the highly conserved domain II of Aux/IAAs abolish the TIR1-Aux/IAA interaction and thus cause an auxin-resistant phenotype. Here we show that peptidyl-prolyl isomerization of rice OsIAA11 catalysed by LATERAL ROOTLESS2 (LRT2), a cyclophilin-type peptidyl-prolyl cis/trans isomerase, directly regulates the stability of OsIAA11. NMR spectroscopy reveals that LRT2 efficiently catalyses the cis/trans isomerization of OsIAA11. The lrt2 mutation reduces OsTIR1-OsIAA11 interaction and consequently causes the accumulation of a higher level of OsIAA11 protein. Moreover, knockdown of the OsIAA11 expression partially rescues the lrt2 mutant phenotype in lateral root development. Together, these results illustrate cyclophilincatalysed peptidyl-prolyl isomerization promotes Aux/IAA degradation, as a mechanism regulating auxin signalling.

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Tuning a timing device that regulates lateral root development in rice

et al. 2019

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Peptidyl Prolyl Isomerases (PPIases) accelerate cis-trans isomerization of prolyl peptide bonds. In rice, the PPIase LRT2 is essential for lateral root initiation. LRT2 displays in vitro isomerization of a highly conserved W-P peptide bond ( 104 W-P 105 ) in the natural substrate OsIAA11. OsIAA11 is a transcription repressor that, in response to the plant hormone auxin, is targeted to ubiquitinmediated proteasomal degradation via specific recognition of the cis isomer of its 104 W-P 105 peptide bond. OsIAA11 controls transcription of specific genes, including its own, that are required for lateral root development. This auxin-responsive negative feedback circuit governs patterning and development of lateral roots along the primary root. The ability to tune LRT2 activity via mutagenesis is crucial for understanding and modeling the role of this bimodal switch in the auxin circuit and lateral root development. We present characterization of the thermal stability and isomerization rates of several LRT2 mutants acting on the OsIAA11 substrate. The thermally stable mutants display activities lower than that of wild-type (WT) LRT2. These include binding diminished but catalytically active P125K, binding incompetent W128A, and binding capable but catalytically incompetent H133Q mutations. Additionally, LRT2 homologs hCypA from human, TaCypA from Triticum aestivum (wheat) and PPIB from E. coli were shown to have 110%, 50% and 60% of WT LRT2 activity on the OsIAA11 substrate. These studies identify several thermally stable LRT2 mutants with altered activities that will be useful for establishing relationships between cis-trans isomerization, auxin circuit dynamics, and lateral root development in rice.

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Structural and biochemical characterization of the cytosolic wheat cyclophilin TaCypA-1

Cited by 17 publications

References 42 publications

A Redox 2-Cys Mechanism Regulates the Catalytic Activity of Divergent Cyclophilins

A Redox 2-Cys Mechanism Regulates the Catalytic Activity of Divergent Cyclophilins

Peptidyl-prolyl isomerization targets rice Aux/IAAs for proteasomal degradation during auxin signalling

Tuning a timing device that regulates lateral root development in rice

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