Abstract

Protein-protein interactions between the C-terminal domain of Myosin Binding Subunit (MBS) of MLC Phosphatase (MBS_CT180; C-terminal 180 aa) and the N-terminal coiled coil (CC) leucine zipper (LZ) domain of PKGI α, PKG-Iα_1-159 play an important role in the process of Smooth Muscle Cell relaxation. The paucity of three-dimensional structural information for MBSCT180 prevents an atomic level understanding of the MBS–PKG contractile complex. MBS_CT180 is comprised of three structurally different sub-domains including a non-canonical CC, a CC, and a LZ. Recently we reported polypeptide purification and biophysical characterization of the CC domain and the LZ domain of MBS_CT180 (Sharma et al, Prot Expr Purif 2012). Here we report ¹H, ¹³C, ¹⁵N chemical shift assignments of homodimeric CC MBS domain encompassing amino acid residues Asp931-Leu980 using 2D and 3D heteronuclear NMR spectroscopy. Secondary structure analyses deduced from these NMR chemical shift data have identified a contiguous stretch of 36 residues from Phe932 to Ala967 that is involved in the formation of coiled coil α-helical region within CC MBS domain. The N-terminal residue Asp931 and the C-terminally positioned residues Thr968-Ala975, Arg977, and Ser978 adopt nonhelical loop conformations.

Keywords: 2D ¹H-¹⁵N HSQC, Coiled coil MBS, Leucine zipper PKG-Iα_1-159, MLCP, NMR assignment, Secondary structure, VSMC.