Atomic Structures of the Human Immunophilin FKBP-12 Complexes with FK506 and Rapamycin

Duyne, Gregory D. Van; Standaert, Robert F.; Karplus, P. Andrew; Schreiber, Stuart L.; Clardy, Jon

doi:10.1006/jmbi.1993.1012

Cited by 1,161 publications

(929 citation statements)

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“…In all three cases, a complete dataset to 1.7 Å resolution was collected from a single crystal cooled to 100 K using a 345 mm Mar Research image plate detector, with in-house monochromatic Cu KR X-rays generated by a Elliot GX13 rotating anode. This crystal form is isomorphous with that of the published FKBP12-rapamycin complex (van Duyne et al, 1991a(van Duyne et al, , 1991b(van Duyne et al, , 1993, with the notable distinction that for both mutants there is a 3.4 Å reduction in the unit cell c axis (due to a rigid body shift of the protein molecule: see below). This is a direct result of the structural response to mutation, discussed below.…”

Section: Expression and Purification Of The Wild-type And Mutantsmentioning

confidence: 99%

Energetic and Structural Analysis of the Role of Tryptophan 59 in FKBP12

Fulton¹,

Jackson

Buckle

2003

Biochemistry

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Tryptophan 59 forms the seat of the hydrophobic ligand-binding site in the small immunophilin FKBP12. Mutating this residue to phenylalanine or leucine stabilizes the protein by 2.72 and 2.35 kcal mol -1 , respectively. Here we report the stability data and 1.7 Å resolution crystal structures of both mutant proteins, complexed with the immunosuppressant rapamycin. Both structures show a relatively large response to mutation involving a helical bulge at the mutation site and the loss of a hydrogen bond that anchors a nearby loop. The increased stability of the mutants is probably due to a combination of improved packing and an entropic gain at the mutation site. The structures are almost identical to that of wild-type FKBP12.6, an isoform of FKBP12 that differs by 18 residues, including Trp59, in its sequence. Therefore, the structural difference between the two isoforms can be attributed almost entirely to the identity of residue 59. It is likely that in FKBP12-ligand complexes Trp59 provides added binding energy at the active site at the expense of protein stability, a characteristic common to other proteins. FKBP12 associates with the ryanodine receptor in skeletal muscle (RyR1), while FKBP12.6 selectively binds the ryanodine receptor in cardiac muscle (RyR2). The structural response to mutation suggests that residue 59 contributes to the specificity of binding between FKBP12 isoforms and ryanodine receptors.

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Section: Expression and Purification Of The Wild-type And Mutantsmentioning

confidence: 99%

Energetic and Structural Analysis of the Role of Tryptophan 59 in FKBP12

Fulton¹,

Jackson

Buckle

2003

Biochemistry

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“…Selenomethionine TTPARG was produced in the same Rosetta (DE3) cells as native TTPARG, but in the conditions where the methionine biosynthesis pathway is inhibited 20 . Briefly, 1 l of unlabelled TBGG ('Terrific Broth' supplemented with 1.0% glucose and 4% glycerol) was inoculated with 5 ml of an overnight culture grown in lysogeny broth (LB).…”

Section: Parg Inhibition Assaymentioning

confidence: 99%

Structure and mechanism of a canonical poly(ADP-ribose) glycohydrolase

Dunstan¹,

Barkauskaite

Lafite

et al. 2012

Nat Commun

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Poly(ADP-ribosyl)ation is a reversible post-translational protein modification involved in the regulation of a number of cellular processes including DnA repair, chromatin structure, mitosis, transcription, checkpoint activation, apoptosis and asexual development. The reversion of poly(ADP-ribosyl)ation is catalysed by poly(ADP-ribose) (PAR) glycohydrolase (PARG), which specifically targets the unique PAR (1′′-2′) ribose-ribose bonds. Here we report the structure and mechanism of the first canonical PARG from the protozoan Tetrahymena thermophila. In addition, we reveal the structure of T. thermophila PARG in a complex with a novel rhodaninecontaining mammalian PARG inhibitor RBPI-3. our data demonstrate that the protozoan PARG represents a good model for human PARG and is therefore likely to prove useful in guiding structure-based discovery of new classes of PARG inhibitors.

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“…The amino acyl residues enclosed to the PPIase active fragment are underlined. Asterisks mark residues of hFKBP12 which are involved in FK506 binding [39]. Bold letters represent residues which are perfectly conserved among FKBPs [39].…”

Section: Enzymatic and Structural Propertiesmentioning

confidence: 99%

An 11.8 kDa proteolytic fragment of the E. coli trigger factor represents the domain carrying the peptidyl‐prolyl cis/trans isomerase activity

Stoller¹,

Tradler²,

Rücknagel³

et al. 1996

FEBS Letters

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The 48 kDa trigger factor (TF) of E. coli was shown to be a peptidyl-prolyl cisltrans isomerase (PPIase). Its location on a ribosomal particle is unique among the PPIases described so far, and suggests a role in de novo protein folding. The trigger factor was investigated with regard to a domain carrying the catalytic activity. An enzymatieally active fragment could be isolated after proteolysis by subtifisin. The resulting polypeptide was analysed by N-terminal sequencing and MALDI-TOF mass spectrometry revealing an 11.8 kDa fragment of TF encompassing the amino acid residues Arg-145 to Glu-251. The nucleotide sequence encoding the amino acid residues Met-140 to Ala-250 of the TF was cloned into vector pQE32. After expression in E. coli the resulting His-tagged polypeptide was isolated on an Ni 2+-NTA column. Subsequent digestion with subtifisin and anionexchange chromatography yielded a TF fragment encompassing amino acids Gin-148 to Thr-249. This fragment may represent the catalytic core of TF since PPIase activity with a specificity constant kcat/Km of 1.3 ~1-1 s -l could be demonstrated when using Suc-Ala-Phe-Pro-Phe-NH-Np as a substrate. Moreover, as was observed for the complete, authentic TF the PPIase activity of the fragment was not inhibited by the peptidomacrofide FK506.

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Atomic Structures of the Human Immunophilin FKBP-12 Complexes with FK506 and Rapamycin

Cited by 1,161 publications

References 0 publications

Energetic and Structural Analysis of the Role of Tryptophan 59 in FKBP12

Energetic and Structural Analysis of the Role of Tryptophan 59 in FKBP12

Structure and mechanism of a canonical poly(ADP-ribose) glycohydrolase

An 11.8 kDa proteolytic fragment of the E. coli trigger factor represents the domain carrying the peptidyl‐prolyl cis/trans isomerase activity

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