NMR experiments on the transient interaction of the intrinsically disordered N-terminal peptide of cystathionine-β-synthase with heme

13C Direct Detected NMR for Challenging Systems

“…Other motifs in which prolines are flanked by specific amino acids are also emerging, as described above for the interaction of α-synuclein with calcium ions through the DPD or EPE motifs. Similar motifs were also identified in different proteins, such as viral proteins ,− as well as in WASP-interacting protein (WIP), an intrinsically disordered polypeptide with a key role in actin polymerization in activated T cells. , Another interesting example in this context is provided by aromatic-proline pairs as recently monitored in different proteins ,, and illustrated below through the example of quail osteopontin . The 13 C detected experiments used to complete resonance assignment revealed in addition to the major form, also a subset of peaks with lower intensity that could be clearly identified in the clean region of CON spectra reporting the correlation of proline nitrogen (Figure ).…”

Cystathionine-β-synthase: Molecular Regulation and Pharmacological Inhibition

“…As for the role of the heme, its function remains vague since it is not directly involved in the catalysis, but still influences folding and is sensitive to the redox status of its environment. In addition, recent studies suggest that the first 40 residues of the human CBS N-terminus constitute an intrinsically disordered region, which transiently binds heme via a second binding site, the CP-based motif with Cys15 and His22 as axial ligands [18,19]. While the function of this additional heme-binding site is not fully understood, according to one publication, the CBS Cys15Ser mutant is unable to bind heme at this second binding site and is ~30% less active compared to the WT variant [18].…”

“…Together, these residues anchor the PLP deeply in the protein matrix and the active site is accessible only through a narrow channel. Conformation of the loops delineating the PLP-containing cavity, namely L145-148, L171-174, and L191-202, defines accessibility of the catalytic center by the substrates and thus the activity of the enzyme (Figure 4) [19,194].…”

“…SAM is a V-type activator of CBS meaning that it increases catalytic efficiency by increasing Vmax of CBS without any significant effect on affinity of the substrates (km). Crystal structures of human CBS in both the SAM-free basal and the SAM-bound activated conformations provided further insight into molecular mechanism of the allosteric regulation of CBS by SAM (Figure 7) [19,195,196].…”

“…Therefore, each CBS subunit contains two potential SAM bindings sites. However, crystal structures of human CBS showed that only one site can accommodate SAM, while the other site is blocked by several bulky hydrophobic residues (Figure 7) [19,195,196]. Binding of SAM into the only available site induces rotation of the CBS domains, which weakens their interactions with the loops of the catalytic cavity.…”