Methionin spin-labeled

Clearly, the potential to use spin labels as a means to reduce protein concentration for detection of protein-ligand interactions, given by the factor of 6582, is tremendous. The sixth-power dependence on electron-proton distance underlines the need to carefully design the residue type which is to be spin labeled. Residues that can be spin labeled include lysine, tyrosine, cysteine, histidine, and methionine [7, 14]. At least one residue of... 

Very fast electron transfers from P+ to bacteriochlorophyl (Bchl) and from (Bchl)- to QA do not depend on media dynamics and occur via conformationally non-equilibrium states (Fig.3.18). The dual fluorophore-nitroxide molecules (D-A) are also convenient objects for analysing the activity-dynamics relationship. The marked irreversible photoreduction of the nitroxide fragment of the dual probe incorporated into the binding site of HSA only took place when the nanosecond dynamical processes around the probe traced by ESR and fluorescence methods were detected (Rubtsova et al., 1993, Fogel et al, 1994 Likhtenshtein, 1986 Lozinsky et al., 2002). Similar results were reported for another model protein system, i.e. a-chymotrypsin with spin labeled methionin-92 groups (Belonogova et al., 1997). In the latter enzyme, the excited tryptophan group serves as an electron donor. 

Muscle contraction is regulated, to a significant extent, by the interaction of troponin (Tn) proteins C, I and T. The interactions between Tn I and Tn C were studied by NMR using Tn C with a spin label attached to Cys84 and specifically 13C-labelled at its 10 methionine residues.208 From the known assignments of the methionine methyl cross-peaks in the HSQC spectrum of Tn C, and their absence from HSQC spectra in the presence of fragments of the bound Tn I, it was concluded that the linker between the two domains of Tn C is flexible, allowing association between the two domains in the presence of Tn 1. 

Figure 15.2-9 shows an example of selectively 15N-Met labeled kinase BTK, which was expressed in baculovirus-infected insect cells. For this kinase no assignment is available so far. The [ H,15N]-TROSY spectra shows 12 peaks corresponding to the 12 methionines of the primary sequence (Fig. 15.2-7(a)). Upon adding the spin-labeled adenosine, four peaks are strongly attenuated corresponding to the four methionines which are in close distance to the spin label. This information is very valuable for the assignment process. 

Spin labels are stable, paramagnetic molecules that, by their structme, easily attach themselves to various biological macromolecular systems such as proteins or cell membranes. Examples of spin labels that can be covalently bonded to specific sites of biological systems include nitroxide derivatives of A-ethylmaleimide, which bind specifically to -SH groups, and nitroxide derivatives of iodoacetamide, which bind specifically to methionine, lysine, and arginine residues of amino acids. Nonco-valently bonded spin-labels that can be incorporated into biological systems include nitroxide derivatives of stearic acid, of phospholipids, and of cholesterol. 

---