RESUMO
The c-MYC transcription factor is a master regulator of cell growth and proliferation and is an established target for cancer therapy. This basic helix-loop-helix Zip protein forms a heterodimer with its obligatory partner MAX, which binds to DNA via the basic region. Considerable research efforts are focused on targeting the heterodimerization interface and the interaction of the complex with DNA. The only available crystal structure is that of a c-MYC:MAX complex artificially tethered by an engineered disulfide linker and prebound to DNA. We have carried out a detailed structural analysis of the apo form of the c-MYC:MAX complex, with no artificial linker, both in solution using nuclear magnetic resonance (NMR) spectroscopy and by X-ray crystallography. We have obtained crystal structures in three different crystal forms, with resolutions between 1.35 and 2.2 Å, that show extensive helical structure in the basic region. Determination of the α-helical propensity using NMR chemical shift analysis shows that the basic region of c-MYC and, to a lesser extent, that of MAX populate helical conformations. We have also assigned the NMR spectra of the c-MYC basic helix-loop-helix Zip motif in the absence of MAX and showed that the basic region has an intrinsic helical propensity even in the absence of its dimerization partner. The presence of helical structure in the basic regions in the absence of DNA suggests that the molecular recognition occurs via a conformational selection rather than an induced fit. Our work provides both insight into the mechanism of DNA binding and structural information to aid in the development of MYC inhibitors.
Assuntos
Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/química , Cristalografia por Raios X/métodos , Proteínas de Ligação a DNA/química , DNA/química , Sequências Hélice-Alça-Hélice/fisiologia , Espectroscopia de Ressonância Magnética/métodos , Proteínas Repressoras/química , Fatores de Transcrição/química , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Galinhas , DNA/genética , DNA/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Humanos , Estrutura Secundária de Proteína , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
c-MYC and the SWI/SNF chromatin remodeling complex act as master regulators of transcription, and play a key role in human cancer. Although they are known to interact, the molecular details of their interaction are lacking. We have determined the structure of the RPT1 region of the INI1/hSNF5/BAF47/SMARCB1 subunit of the SWI/SNF complex that acts as a c-MYC-binding domain, and have localized the interaction regions on both INI1 and on the c-MYC:MAX heterodimer. c-MYC interacts with a highly conserved groove on INI1, while INI1 binds to the c-MYC helix-loop-helix region. The binding site overlaps with the c-MYC DNA-binding region, and we show that binding of INI1 and E-box DNA to c-MYC:MAX are mutually exclusive.
Assuntos
Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/química , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Proteínas Cromossômicas não Histona , Proteínas Proto-Oncogênicas c-myc/química , Proteínas Proto-Oncogênicas c-myc/metabolismo , Proteína SMARCB1/química , Proteína SMARCB1/metabolismo , Fatores de Transcrição , Sequência de Aminoácidos , Sítios de Ligação , Cristalografia por Raios X , Humanos , Modelos Moleculares , Conformação Proteica , Domínios Proteicos , Multimerização ProteicaRESUMO
In this review we discuss recent progress in targeting the protein-protein interactions made by oncogenic transcription factors. We particularly focus on the challenges posed by the prevalence of intrinsically disordered regions in this class of protein and the strategies being used to overcome them.
Assuntos
Descoberta de Drogas/métodos , Proteínas Intrinsicamente Desordenadas/química , Proteínas Intrinsicamente Desordenadas/metabolismo , Fatores de Transcrição/química , Fatores de Transcrição/metabolismo , Animais , Humanos , Ligação Proteica/efeitos dos fármacosRESUMO
Thymidylate synthase (TS) is a target for antifolate-based chemotherapies of microbial and human diseases. Here, ligand-based, synthetic, and X-ray crystallography studies led to the discovery of 6-(3-cyanobenzoyloxy)-2-oxo-2H-naphto[1,8-bc]furan, a novel inhibitor with a Ki of 310 nM against Pneumocystis carinii TS. The X-ray ternary complex with Escherichia coli TS revealed, for the first time, displacement of the substrate toward the dimeric protein interface, thus providing new opportunities for further design of specific inhibitors of microbial pathogens.