SH2-Catalytic Domain Linker Heterogeneity Influences
Allosteric Coupling across the SFK Family
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Abstract
Src-family
kinases (SFKs) make up a family of nine homologous multidomain
tyrosine kinases whose misregulation is responsible for human disease
(cancer, diabetes, inflammation, etc.). Despite overall sequence homology
and identical domain architecture, differences in SH3 and SH2 regulatory
domain accessibility and ability to allosterically autoinhibit the
ATP-binding site have been observed for the prototypical SFKs Src
and Hck. Biochemical and structural studies indicate that the SH2-catalytic
domain (SH2-CD) linker, the intramolecular binding epitope for SFK
SH3 domains, is responsible for allosterically coupling SH3 domain
engagement to autoinhibition of the ATP-binding site through the conformation
of the αC helix. As a relatively unconserved region between
SFK family members, SH2-CD linker sequence variability across the
SFK family is likely a source of nonredundant cellular functions between
individual SFKs via its effect on the availability of SH3 and SH2
domains for intermolecular interactions and post-translational modification.
Using a combination of SFKs engineered with enhanced or weakened regulatory
domain intramolecular interactions and conformation-selective inhibitors
that report αC helix conformation, this study explores how SH2-CD
sequence heterogeneity affects allosteric coupling across the SFK
family by examining Lyn, Fyn1, and Fyn2. Analyses of Fyn1 and Fyn2,
isoforms that are identical but for a 50-residue sequence spanning
the SH2-CD linker, demonstrate that SH2-CD linker sequence differences
can have profound effects on allosteric coupling between otherwise
identical kinases. Most notably, a dampened allosteric connection
between the SH3 domain and αC helix leads to greater autoinhibitory
phosphorylation by Csk, illustrating the complex effects of SH2-CD
linker sequence on cellular function