Table 1. Design of the small focused N-TIMP-2 library.

Desired amino acid choices display mutations that were tested experimentally and showed enhanced binding specificity of N-TIMP-2 towards MMP-14_CAT in our previous work when introduced as single mutations. The rightmost column shows all the amino acid choices encoded in the library at each position, including extra choices that appeared due to encoding each position with a particular degenerate codon. WT amino acid identities are underlined.

N-TIMP-2 Position	Desired Amino Acid Choices	Degenerate Codon	Actual Encoded Amino Acids
4	A, Q, E, S	BMR	A, Q, E, P, S, stop
6	R, V	SKN	R, G, L, V
35	E, I, K	RWA	E, I, K, V
38	N, Q	MAN	N, Q, H, K
68	S, W, Y	WRS	R, N, C, K, S, W, Y, stop
71	N, V	RWY	N, D, I, V
97	R, H	CRY	R, H
99	T, Y	WMY	N, S, T, Y