Interactions with the Bifunctional Interface of the Transcriptional Coactivator DCoH1 Are Kinetically Regulated

• Published in: The Journal of biological chemistry Vol. 290; no. 7; pp. 4319 - 4329
• Main Authors: Wang, Dongli, Coco, Matthew W., Rose, Robert B.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 13.02.2015; American Society for Biochemistry and Molecular Biology
• Subjects: Animals; BASIC BIOLOGICAL SCIENCES; Chromatography, Gel; Crystallography, X-Ray; DCoH1; Hydro-Lyases - chemistry; Hydro-Lyases - genetics; Hydro-Lyases - metabolism; INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY; Kinetic Hot Spot; Kinetic Stability; Kinetics; Mice; Models, Molecular; MODY; Multifunctional Protein; Mutation - genetics; PCD; Protein Complex; Protein Conformation; Protein Folding; Protein Multimerization; Protein Structure and Folding; Serine - chemistry; Serine - genetics; Serine - metabolism; Threonine - chemistry; Threonine - genetics; Threonine - metabolism; Transcriptional Coactivator; Water - chemistry; PCD; Transcriptional Coactivator; Kinetic Hot Spot; MODY; DCoH1; Kinetic Stability; Protein Complex; Kinetics; Protein Folding; Multifunctional Protein
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M114.616870

Pterin-4a-carbinolamine dehydratase (PCD) is a highly conserved enzyme that evolved a second, unrelated function in mammals, as a transcriptional coactivator. As a coactivator, PCD is known as DCoH or dimerization cofactor of the transcription factor HNF-1. These two activities are associated with a change in oligomeric state: from two dimers interacting as an enzyme in the cytoplasm to a dimer interacting with a dimer of HNF-1 in the nucleus. The same interface of DCoH forms both complexes. To determine how DCoH partitions between its two functions, we studied the folding and stability of the DCoH homotetramer. We show that the DCoH1 homotetramer is kinetically trapped, meaning once it forms it will not dissociate to interact with HNF-1. In contrast, DCoH2, a paralog of DCoH1, unfolds within hours. A simple mutation in the interface of DCoH2 from Ser-51 to Thr, as found in DCoH1, increases the kinetic stability by 9 orders of magnitude, to τ½ ∼2 million years. This suggests that the DCoH1·HNF-1 complex must co-fold to interact. We conclude that simple mutations can dramatically affect the dissociation kinetics of a complex. Residue 51 represents a “kinetic hot spot” instead of a “thermodynamic hot spot.” Kinetic regulation allows PCD to adopt two distinct functions. Mutations in DCoH1 associated with diabetes affect both functions of DCoH1, perhaps by disrupting the balance between the two DCoH complexes. Background: The protein dimerization cofactor of HNF-1 (DCoH1)/pterin-carbinolamine dehydratase (PCD) accomplishes two unrelated activities, forming two complexes with the same interface. Results: The DCoH1 homotetramer is kinetically trapped; a single mutation in the interface increases the unfolding rate 109-fold. Conclusion: Kinetic regulation allows DCoH to assume two unrelated functions. Significance: Mutations excluding water from interfaces represent kinetic “hot spots,” dramatically affecting dissociation rates.