Design, synthesis, and kinetic evaluation of high-affinity FKBP ligands and the X-ray crystal structures of their complexes with FKBP12

Holt, Dennis A.; Luengo, Juan I.; Yamashita, Dennis S.; Oh, H.-J.; Konialian, Arda L.; Yen, Hwa Kwo; Rozamus, Leonard W.; Brandt, Martin S.; Bossard, Mary J.; Levy, Mark A.; Eggleston, Drake S.; Liang, Jun; Schultz, L. Wayne; Stout, Thomas J.; Clardy, Jon

doi:10.1021/ja00075a008

Cited by 219 publications

(326 citation statements)

References 11 publications

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“…Both molecules are capable of simultaneously binding to FKBP and the Fyn SH2 domain. The pYEEI peptide was linked covalently to two FKBP ligands, FK506 and SLF (15), to provide the desired bifunctional molecules, FKpYEEI and SLFpYEEI (Fig. 1).…”

Section: Resultsmentioning

confidence: 99%

“…SLF is smaller than FK506 and does not project as far from the FKBP protein surface. By using the three-dimensional structures of FKBP12 (15,18), FKBP52 (19), and the Fyn SH2 domain (20), the linkers between the two halves of the bifunctional molecules were designed to bring the FKBP surface into close proximity to the SH2 domain surface. The affinities of FKpYEEI and SLFpYEEI for recombinant FKBP12, FKBP52, and the Fyn SH2 domain were measured by using isothermal titration calorimetry (ITC, Table 1).…”

Section: Resultsmentioning

confidence: 99%

“…The coupled product was treated with hydrogen fluoride in acetonitrile to remove the two silyl ether protecting groups, and the desired product was purified by using reverse-phase HPLC. Synthetic ligand for FKBP (SLF) was synthesized according to the procedures of Holt et al (15). SLF was coupled to the N terminus of the resin-bound protected pYEEI peptide by using PyBOP.…”

Section: Methodsmentioning

confidence: 99%

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Affinity modulation of small-molecule ligands by borrowing endogenous protein surfaces

Briesewitz

Ray

Wandless

et al. 1999

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

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A general strategy is described for improving the binding properties of small-molecule ligands to protein targets. A bifunctional molecule is created by chemically linking a ligand of interest to another small molecule that binds tightly to a second protein. When the ligand of interest is presented to the target protein by the second protein, additional proteinprotein interactions outside of the ligand-binding sites serve either to increase or decrease the affinity of the binding event. We have applied this approach to an intractable target, the SH2 domain, and demonstrate a 3-fold enhancement over the natural peptide. This approach provides a way to modulate the potency and specificity of biologically active compounds.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Affinity modulation of small-molecule ligands by borrowing endogenous protein surfaces

Briesewitz

Ray

Wandless

et al. 1999

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

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“…In order to find novel inhibitors of Mip, two previously reported pipecoline ring containing inhibitors A and B with K i (app) of 10 nM and 0.23 µM, respectively, were used as lead structures. While structure A contained the keto amide moiety, in structure B this group was replaced by a sulfonamide anchor (Holt et al, 1993(Holt et al, , 1994. Molecular docking studies revealed that structure A fitted less well to Mip than to the human homolog FKBP12.…”

Section: Pipecolinic Acid Derivativesmentioning

confidence: 99%

FKBPs in bacterial infections

Ünal

Steinert

2015

Biochimica et Biophysica Acta (BBA) - General Subjects

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“…This compound binds to FKBP but, unlike rapamycin, does not possess affinity for FRB (Holt et al, 1993); thus, it can be used to compete for binding to FKBP and, perhaps, accelerate dissociation of the ternary FKBP-rapamycin-FRB complex. To test whether this competitor can influence the kinetics of protein localization, SLF (10 µM) was added to the culture medium after removal of rapamycin-containing medium.…”

Section: Reversibility Of Drug-based Nuclear Import and Exportmentioning

confidence: 99%

A small molecule‐directed approach to control protein localization and function

Geda

Patury

et al. 2008

Yeast

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Protein localization is tightly linked with function, such that the subcellular distribution of a protein serves as an important control point regulating activity. Exploiting this regulatory mechanism, we present here a general approach by which protein location, and hence function, may be controlled on demand in the budding yeast. In this system a small molecule, rapamycin, is used to temporarily recruit a strong cellular address signal to the target protein, placing subcellular localization under control of the selective chemical stimulus. The kinetics of this system are rapid: rapamycin-directed nucleo-cytoplasmic transport is evident 10-12 min posttreatment and the process is reversible upon removal of rapamycin. Accordingly, we envision this platform as a promising approach for the systematic construction of conditional loss-of-function mutants. As proof of principle, we used this system to direct nuclear export of the essential heat shock transcription factor Hsf1p, thereby mimicking the cell-cycle arrest phenotype of an hsf1 temperature-sensitive mutant. Our drug-induced localization platform also provides a method by which protein localization can be uncoupled from endogenous cell signalling events, addressing the necessity or sufficiency of a given localization shift for a particular cell process. To illustrate, we directed the nuclear import of the calcineurin-dependent transcription factor Crz1p in the absence of native stimuli; this analysis directly substantiates that nuclear translocation of this protein is insufficient for its transcriptional activity. In total, this technology represents a powerful method for the generation of conditional alleles and directed mislocalization studies in yeast, with potential applicability on a genome-wide scale.

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Design, synthesis, and kinetic evaluation of high-affinity FKBP ligands and the X-ray crystal structures of their complexes with FKBP12

Cited by 219 publications

References 11 publications

Affinity modulation of small-molecule ligands by borrowing endogenous protein surfaces

Affinity modulation of small-molecule ligands by borrowing endogenous protein surfaces

FKBPs in bacterial infections

A small molecule‐directed approach to control protein localization and function

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