Exploring Enzyme Catalysis: A 3D Model of the Myocobacterium Tuberculosis MazF‐mt6 Toxin

Toxin‐antitoxin systems are present in bacterial and archaeal genomes and regulate cellular survival in response to stressors including antibiotics and host immune responses. The toxin‐antitoxin systems all consist of a toxin protein that blocks essential cellular functions such as translation, and either an antitoxin RNA or protein that inhibits toxin function through a variety of mechanisms. Escherichia coli has at least 36 assumed toxin‐antitoxin gene pairs, whereas the pathogenic Myocobacterium tuberculosis has over 90 toxin‐antitoxin pairs. The E. coli MazF toxin proteins are ribonucleases that cleave cellular mRNA once stress has been encountered, while the nine M. tuberculosis MazF family proteins cleave mRNA, tRNA, or rRNA. MazF‐mt6 cleaves helix 70 of the 23S rRNA. Here, we have designed a 3D of model of the MazF‐mt6 protein using Jmol that highlights the overall architecture and the active site based on the crystal structure solved by Dr. Dunham and colleagues (PDB 5UCT; Hoffer et al. 2017 JBC 292 (19):7718–26). MazF‐mt6 protein has Pemk‐like fold with three alpha‐helices, shown in grey, and 2 beta‐sheets shown in purple, one of which folds into a five‐stranded beta‐barrel fold similar to the SH3 domain. The active site is solvent accessible and contains several charged or polar residues that may be involved in catalysis. Our 3D model highlights the critical residues in the active site. Site‐directed mutagenesis studies reveal that Asp10, Arg13, Lys11, and Thr36 are necessary for MazF‐mt6 induced growth arrest, whereas Thr37 and Thr38 are not. In addition, Asp10, Arg13, and Thr36 are necessary for enzymatic activity, suggesting that they are part of a catalytic triad. Designing 3D protein models enables students to explore complex molecular structure‐function relationships.Support or Funding InformationNSF DUE1725940 and NSF HRD1623340This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.