Proteins are an important part of life, of which enzymes perform vital roles in biological systems. Rational protein design is a key approach for investigating the structure and function relationship of enzymes. This review summarizes the progresses in rational design and functional study of artificial hydrolases in protein scaffolds, including reuse of natural proteins, reconstruction of natural proteins, molecular design based on 3-α-helix, 4-α-helix or zinc-finger proteins, and fine-tuning the hydrolysis activity of heme proteins such as cytochrome b₅ and myoglobin mutants. It illustrates the basic idea and approaches of artificial hydrolase design, which provides valuable clues for rational construction of artificial hydrolase or other enzymes. The progress in rational design of artificial hydrolases not only enriches our understanding of the structure-property-reactivity-function relationship of native enzymes, but also enhances our ability to construct artificial enzymes with advanced functions.

Contents
1 Introduction
2 Mechanisms of hydrolases
3 Hydrolases containing no metal ions
3.1 Reuse of natural proteins
3.2 Reconstruction of natural proteins
4 Hydrolases containing metal ions
4.1 Molecular design based on 3-α-helix
4.2 Molecular design based on 4-α-helix
4.3 Molecular design based on zinc-finger protein
5 Hydrolases containing metal complex
5.1 Explore hydrolysis activity of heme proteins
5.2 Tune hydrolysis activity of heme proteins
6 Conclusion and outlook