title: Distal loop flexibility of a regulatory domain modulates dynamics and activity of C-terminal SRC kinase (csk).
abstract: The Src family of tyrosine kinases (SFKs) regulate numerous aspects of cell growth and differentiation and are under the principal control of the C-terminal Src Kinase (Csk). Csk and SFKs share a modular design with the kinase domain downstream of the N-terminal SH2 and SH3 domains that regulate catalytic function and membrane localization. While the function of interfacial segments in these multidomain kinases are well-investigated, little is known about how surface sites and long-range, allosteric coupling control protein dynamics and catalytic function. The SH2 domain of Csk is an essential component for the down-regulation of all SFKs. A unique feature of the SH2 domain of Csk is the tight turn in place of the canonical CD loop in a surface site far removed from kinase domain interactions. In this study, we used a combination of experimental and computational methods to probe the importance of this difference by constructing a Csk variant with a longer SH2 CD loop to mimic the flexibility found in homologous kinase SH2 domains. Our results indicate that while the fold and function of the isolated domain and the full-length kinase are not affected by loop elongation, native protein dynamics that are essential for efficient catalysis are perturbed. We also identify key motifs and routes through which the distal SH2 site might influence catalysis at the active site. This study underscores the sensitivity of intramolecular signaling and catalysis to native protein dynamics that arise from modest changes in allosteric regions while providing a potential strategy to alter intrinsic activity and signaling modulation.
authors.affiliation: Department of Chemistry and Biochemistry, University of California at San Diego, La Jolla, California, United States of America.
authors.forename: Sulyman
authors.initials: S
authors.lastname: Barkho
authors.forename: Levi C T
authors.initials: LC
authors.lastname: Pierce
authors.forename: Maria L
authors.initials: ML
authors.lastname: McGlone
authors.forename: Sheng
authors.initials: S
authors.lastname: Li
authors.forename: Virgil L
authors.initials: VL
authors.lastname: Woods
authors.forename: Ross C
authors.initials: RC
authors.lastname: Walker
authors.forename: Joseph A
authors.initials: JA
authors.lastname: Adams
authors.forename: Patricia A
authors.initials: PA
authors.lastname: Jennings
mesh_terms: D000595:Amino Acid Sequence; D055162:Biocatalysis; D013058:Mass Spectrometry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D017386:Sequence Homology, Amino Acid; D018909:src Homology Domains; D019061:src-Family Kinases / Q000737:chemistry / Q000378:metabolism*
chemical_list: D019061:src-Family Kinases
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