Loop Dynamics of the Extracellular Domain of Human Tissue Factor and Activation of Factor VIIa

Minazzo, Agnese S.; Darlington, Reuben C.; Ross, J. B. Alexander

doi:10.1016/j.bpj.2008.10.018

Cited by 14 publications

(11 citation statements)

References 61 publications

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“…The experiments were performed using time-correlated single-photon counting (TCSPC) as described in ref 51. Steady-state anisotropy values were obtained from the data with the FluoFit Pro version 4.2.1 analysis software package (PicoQuant, Berlin, Germany).…”

Section: Methodsmentioning

confidence: 99%

Structural and Functional Characterization of the Acidic Region from the RIZ Tumor Suppressor

et al. 2015

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RIZ (retinoblastoma protein-interacting zinc finger protein), also denoted PRDM2, is a transcriptional regulator and tumor suppressor. It was initially identified because of its ability to interact with another well-established tumor suppressor, the retinoblastoma protein (Rb). A short motif, IRCDE, in the acidic region (AR) of RIZ was reported to play an important role in the interaction with the pocket domain of Rb. The IRCDE motif is similar to a consensus Rb-binding sequence LXCXE (where X denotes any amino acid) that is found in several viral Rb-inactivating oncoproteins. To improve our understanding of the molecular basis of binding of Rb to RIZ, we investigated the interaction between purified recombinant AR and the pocket domain of Rb using nuclear magnetic resonance spectroscopy, isothermal titration calorimetry, and fluorescence anisotropy experiments. We show that AR is intrinsically disordered and that it binds the pocket domain with submicromolar affinity. We also demonstrate that the interaction between AR and the pocket domain is mediated primarily by the short stretch of residues containing the IRCDE motif and that the contribution of other parts of AR to the interaction with the pocket domain is minimal. Overall, our data provide clear evidence that RIZ is one of the few cellular proteins that can interact directly with the LXCXE-binding cleft on Rb.

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Section: Methodsmentioning

confidence: 99%

Structural and Functional Characterization of the Acidic Region from the RIZ Tumor Suppressor

et al. 2015

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“…The time-dependent anisotropy decays at variable temperature were analyzed using single-exponential correlation times and a nonzero baseline limiting anisotropy ( r ∞ ), which reflects restricted motion of the probe during the lifetime of the excited-state. 32,36-38 …”

Section: Resultsmentioning

confidence: 99%

Photophysical Studies of Bioconjugated Ruthenium Metal–Ligand Complexes Incorporated in Phospholipid Membrane Bilayers

et al. 2013

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Luminescent, mono-diimine, ruthenium complexes, [(H)Ru(CO)(PPh3)2(dcbpy)][PF6] (1, dcbpy = 4,4′-dicarboxy bipyridyl) and [(H)Ru(CO)(dppene)(5-amino-1,10-phen)][PF6] (2, dppene = bis diphenylphosphino-ethylene, phen = 9,10-phenanthroline), have been conjugated with 1,2-dihexadecanoyl-sn-glycero-3-phosphoethanolamine (DPPE) and with cholesterol in the case of 2. Compound 1 gives the bis-lipid derivative [(H)Ru(CO)(PPh3)2(dcbpy-N-DPPE2)][PF6] (3), while 2 provides the mono-lipid conjugate [(H)Ru(CO)(dppene)(1,10-phen-5-NHC(S)-N-DPPE)][ PF6] (4), and the cholesterol derivative [(H)Ru(CO)(dppene)(1,10-phen-5-NHC(O)OChol)][PF6] (5, Chol = cholesteryl), using standard conjugation techniques. These compounds were characterized by spectroscopic methods, and their photophysical properties were measured in organic solvents. The luminescence of lipid conjugates 3 and is quenched in organic solvents while compound 4 a weak, short-lived, blue-shifted emission in solution. The cholesterol conjugate shows the long-lived, microsecond-timescale emission associated with triplet metal-to-ligand charge-transfer (3MLCT) excited states. Incorporation of conjugate 3 in lipid bilayer vesicles restores the luminescence, but with blue shifts (~80 nm) accompanied by nanosecond-timescale lifetimes. In the vesicles conjugate 4 shows a similar short-lived and blue-shifted emission to that observed in solution but with increased intensity. Conjugation of the complex [(H)Ru(CO)(PhP2C2H4C(O)O-N-succinimidyl)2(bpy)][PF6] (6”) with DPPE gives the phosphine-conjugated complex [(H)Ru(CO)(PhP2C2H4C(O)-N-DPPE)2(bpy)][PF6] (7). Complex 7 also exhibits a short-lived and blue-shifted emission in solution and in vesicles as observed for 3 and 4. We have also conjugated the complex [Ru(bpy)2(5-amino-1,10-phenanthroline)][PF6]2 (8) with both cholesterol (9) and DPPE (10). Neither 9 nor the previously reported 10 exhibited the blue shifts observed for 3 and 4 when incorporated into LUVs. The anisotropies of the emissions of 3, 4 and 7 were also measured in LUVs and of 5 in both glycerol and LUVs. High fundamental anisotropies were observed for 3 and 4 and 7.

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“…183,184,190,191 One should nally consider the possibility that usGNP complexation could modulate the conformational entropy/internal dynamics of proteins without major accompanied structural changes. Techniques suitable for probing dynamics include time-resolved uorescence anisotropy, 192,193 NMR spectroscopy 194 and hydrogen/deuterium exchange mass spectrometry. 195 However, the characterization of protein dynamics has remained largely unexplored in usGNP-protein interaction studies.…”

Section: Biomolecular Interactions Of Usgnps With Isolated Proteins In Dilute Solutionsmentioning

confidence: 99%

Biomolecular interactions of ultrasmall metallic nanoparticles and nanoclusters

2021

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Loop Dynamics of the Extracellular Domain of Human Tissue Factor and Activation of Factor VIIa

Cited by 14 publications

References 61 publications

Structural and Functional Characterization of the Acidic Region from the RIZ Tumor Suppressor

Structural and Functional Characterization of the Acidic Region from the RIZ Tumor Suppressor

Photophysical Studies of Bioconjugated Ruthenium Metal–Ligand Complexes Incorporated in Phospholipid Membrane Bilayers

Biomolecular interactions of ultrasmall metallic nanoparticles and nanoclusters

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