Engineering a zinc binding site into the de novo designed protein DS119 with a βαβ structure

Functional proteins designed de novo have potential application in chemical engineering,agriculture and healthcare.Metal binding sites are commonly used to incorporate functions.Based on a de novo designed protein DS119 with aβαβstructure,we have computationally engineered zinc binding sites into it using a home-made searching program.Seven out of the eight designed sequences tested were shown to bind Zn2+with micromolar affinity,and one of them bound Zn2+with 1:1 stoichiometry.This is the first time that metalloproteins with anα,βmixed structure have been designed from scratch.