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Publication Abstract from PubMed

gamma-Hydroxybutyric acid (GHB) analogs are small molecules that bind competitively to a specific cavity in the oligomeric CaMKIIalpha hub domain. Binding affects conformation and stability of the hub domain, which may explain the neuroprotective action of some of these compounds. Here, we describe molecular details of interaction of the larger-type GHB analog 2-(6-(4-chlorophenyl)imidazo[1,2-b]pyridazine-2-yl)acetic acid (PIPA). Like smaller-type analogs, PIPA binding to the CaMKIIalpha hub domain promoted thermal stability. PIPA additionally modulated CaMKIIalpha activity under sub-maximal CaM concentrations and ultimately led to reduced substrate phosphorylation. A high-resolution X-ray crystal structure of a stabilized CaMKIIalpha (6x mutant) hub construct revealed details of the binding mode of PIPA, which involved outward placement of tryptophan 403 (Trp403), a central residue in a flexible loop close to the upper hub cavity. Small-angle X-ray scattering (SAXS) solution structures and mass photometry of the CaMKIIalpha wild-type hub domain in the presence of PIPA revealed a high degree of ordered self-association (stacks of CaMKIIalpha hub domains). This stacking neither occurred with the smaller compound 3-hydroxycyclopent-1-enecarboxylic acid (HOCPCA), nor when Trp403 was replaced with leucine (W403L). Additionally, CaMKIIalpha W403L hub was stabilized to a larger extent by PIPA compared to CaMKIIalpha hub wild type, indicating that loop flexibility is important for holoenzyme stability. Thus, we propose that ligand-induced outward placement of Trp403 by PIPA, which promotes an unforeseen mechanism of hub domain stacking, may be involved in the observed reduction in CaMKIIalpha kinase activity. Altogether, this sheds new light on allosteric regulation of CaMKIIalpha activity via the hub domain.

Ligand-induced CaMKIIalpha hub Trp403 flip, hub domain stacking, and modulation of kinase activity.,Narayanan D, Larsen ASG, Gauger SJ, Adafia R, Hammershoi RB, Hamborg L, Bruus-Jensen J, Griem-Krey N, Gee CL, Frolund B, Stratton MM, Kuriyan J, Kastrup JS, Langkilde AE, Wellendorph P, Solbak SMO Protein Sci. 2024 Oct;33(10):e5152. doi: 10.1002/pro.5152. PMID:39275999^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.