The development of thermostable and solvent-tolerant metalloproteins is a long-sought goal for many applica- tions in synthetic biology and biotechnology. In this work, we were able to engineer a highly thermo- and organic solvent- stable metallo variant of the B1 domain of protein G (GB1) with a tetrahedral zinc-binding site reminiscent of the one of thermolysin. Promising candidates were designed computationally by applying a protocol based on classical and first-princi- ples molecular dynamics simulations in combination with genetic algorithm (GA) optimization. The most promising of the computationally predicted mutants was expressed and structurally characterized and yielded a highly thermostable protein. The experimental results thus confirm the predictive power of the applied computational protein engineering approach for the de novo design of highly stable metalloproteins.