Journal
BIOCHEMISTRY
Volume 44, Issue 4, Pages 1089-1096Publisher
AMER CHEMICAL SOC
DOI: 10.1021/bi0476127
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- NINDS NIH HHS [NS37200] Funding Source: Medline
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Protein kinase C (PKC) isozymes and other receptors of diacylglycerol (DAG) bind to this widespread second messenger through their C-1 domains. These alternative DAG receptors include munc 13-1, a large neuronal protein that is crucial for DAG-dependent augmentation of neurotransmitter release. Whereas the structures of several PKC C-1 domains have been determined and have been shown to require little conformational changes for ligand binding, it is unclear whether the C-1 domains from other DAG receptors contain specific structural features with key functional significance. To gain insight into this question, we have determined the three-dimensional structure in solution of the munc13-1 C-1 domain using NMR spectroscopy. The overall structure includes two beta-sheets, a short C-terminal alpha-helix, and two Zn2+-binding sites, resembling the structures of PKC C-1 domains. However, the munc13-1 C-1 domain exhibits striking structural differences with the PKC C-1 domains in the ligand-binding site. These differences result in occlusion of the binding site of the munc13-1 C-1 domain by a conserved tryptophan side chain that in PKCs adopts a completely different orientation. As a consequence, the munc13-1 C-1 domain requires a considerable conformational change for ligand binding. This structural distinction is expected to decrease the DAG affinity of munc13-1 compared to that of PKCs, and is likely to be critical for munc13-1 function. On the basis of these results, we propose that augmentation of neurotransmitter release may be activated at higher DAG levels than PKCs as a potential mechanism for uncoupling augmentation of release from the multitude of other signaling processes mediated by DAG.
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