SH3 domain of a-spectrin

The Src homology 3 (SH3) domain of a-spectrin was the first SH3 domain structure to be solved (Musacchio et al., 1992). The domain was initially identified as regions of similar sequence found within signaling proteins, such as the Src family of tyrosine kinases, the Crk adaptor... 

NCOCA- and NCACB-type experiments, such as those discussed in Section 2.6, were used to obtain an almost complete set of de novo resonance assignments for the SH3 domain of a-spectrin (62 residues, 7.2 kDa) under MAS conditions. " As discussed above, correlation of these ( N, C) chemical shift assignments with random coil values subsequently showed " that secondary chemical shifts provide a straightforward and powerful instrument with which to monitor secondary structure in solid-phase proteins under MAS conditions. Moreover, these resonance assignments represented the starting point for an investigation of the proton resonance frequencies and for the structural... 

Scheme 8 Synthesis of the SH3 domain of a-spectrin using 4,5,6-trimethoxy-2-mercaptobenzyl ligation auxiliary 33...

Agarwal et al. have compared the internal dynamics of a small globular protein, SH3 domain from a-spectrin, in solution and in a crystalline state. The comparison involved side-chain methyl R relaxation rates that are highly sensitive to local dynamics in the vicinity of the methyl site. To conduct the relaxation measurements, protein samples were prepared using specially labelled a-ketoisovalerate precursors, resulting in selective incorporation of the spin pair in one or both methyl groups of the... 

Sadqi, M., Casares, S., Ahril, M. A., Lopez-Mayorga, O., Conejero-Lara, F., and Freire, E. (1999). The native state conformational ensemble of the SH3 domain from alpha-spectrin. Biochemistry 38, 8899-8906. 

Blanco, F. J., Ortiz, A. R., and Serrano, L. (1997). and NMR assignment and solution structure of the SH3 domain of spectrin Comparison of unrefined and refined structure sets with the crystal structure./. Biomol. NMR 9, 347-357. 

As an alternative means, it is a natural consequence to expect that high-resolution sohd-state NMR could be conveniently utilized to reveal the 3D structure and dynamics of a variety of membrane proteins, because the expected NMR line widths available from sohd-state NMR are not any more influenced by motional fluctuation of proteins under consideration as a whole as encountered in solution NMR. For instance, an attempt was made to determine 3D structure of uniformly C-labeled a-spectrin SH3 domain as a globular protein, based on distance constraints estimated from proton-driven spin-diffusion (PDSD) measure-... 

Another report on NMR studies of perdeuterated proteins by Agarwal et al. has shown that the small amount of residual protonation is sufficient to perform 2D and 3D MAS solid-state NMR experiments. In particular, a HCCH-TOBSY type experiment was suggested, which was successfully employed to assign the methyl resonances in aliphatic side chains in a perdeuterated sample of the SH3 domain of chicken a-spectrin. 

A comprehensive analysis of the dynamics of the SH3 domain of chicken a-spectrin has been presented, based upon Ti and on- and off-resonance 7 ip relaxation times obtained on deuterated samples with a partial back-exchange of labile protons under a variety of the experimental conditions, taking explicitly into account the dipolar order parameters calculated from dipole-dipole couplings. It is demon-... 

X. Periole, M. Vendruscolo, and A. E. Mark, Proteins Struct., Funct., Bioinf., 69, 536 (2007). Molecular Dynamics Simulations from Putative Transition States of a-Spectrin SH3 Domain. 

Fig. 6 A general pulse sequence (a) for spin diffusion or third-spin assisted mixing for polarization transfer between carbons (as shown) or carbon and nitrogen along with examples of homonuclear mixing elements and illustration of activated parts of H-13C-13C spin systems for PDSD, MIRROR, and PAR (b) (taken from [18] with permission), (c) Example of the application of PDSD for structure determination of the a-spectrin SH3 domain (taken from [133] with permission)...

A primary function of the SH3 domains is to form fimctional oligomeric complexes at defined subcellular sites, frequently in cooperation with other modular domains. SH3 domains are foimd in many proteins associated with the cytoskeleton or with the plasma membrane. Examples are the actin binding protein a-spectrin and myosin lb. Furthermore, SH3 interactions are involved in signal transduction in the Ras pathway (see Chapter 9). 

M. E- Noble, A. Musacchia, M. Saraste, S. A. Courmeidge, and R. K. Wirenga. Crystal structure of the SH3 domain in human Fyn comparison of the three-dimensional structures of SH3 domains in tyrosine kinases and spectrin. EMBOJ, 12, 2617-2624, 1993. 

The backbone and side-chain and N signals of a solid 62-residue (U- C, N)-labelled protein containing the a-spectrin SH3 domain were assigned by 2D MAS N- C and dipolar correlation spectroscopy at... 

Fig. 9. Example of secondary chemical shift correlating to secondary structure, illustrated for the case of the a-spectrin SH3 domain.

Fig. 13. Structures of polypeptides determined by MAS solid-state NMR. The PDB file numbers are shown where available, (a) The dipeptide glycyl isoleucine, (b) The tripeptide iV-formyl-L-Met-L-Eeu-L-Phe-OH. (c) The a-spectrin SH3 domain, (d) An 11-amino acid stretch of transthyretin...

Morel B, Casares S, Coneja-o-Lara F (2006) A single mutation induces amyloid aggregation in the alpha-spectrin SH3 domain analysis of the early stages of fibril formation. J Mol Biol 356 453-468... 

In another paper, a 3D NMR based approach for the determination of the fold of moderately sized proteins by solid state MAS NMR is presented and applied to the a-spectrin SH3 domain.This new methodology includes the measure-... 

High-accuracy measurement of H, N DCs in the backbone of a per-deuterated sample of the a-spectrin SH3 domain was performed by means of a 3D DC experiment by introducing phase-inverted CP (CPPI) element in which H and rf fields are simultaneously phase-alternated every 10 ms... 

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