Increase of enzyme activity through specific covalent modification with fragments

TY - JOUR

T1 - Increase of enzyme activity through specific covalent modification with fragments

AU - Darby, John F.

AU - Atobe, Masakazu

AU - Firth, James D.

AU - Bond, Paul

AU - Davies, Gideon J.

AU - O'Brien, Peter Andrew

AU - Hubbard, Roderick E.

N1 - © The Royal Society of Chemistry 2017.

PY - 2017/11/1

Y1 - 2017/11/1

N2 - Modulation of enzyme activity is a powerful means of probing cellular function and can be exploited for diverse applications. Here, we explore a method of enzyme activation where covalent tethering of a small molecule to an enzyme can increase catalytic activity (k cat/K M) up to 35-fold. Using a bacterial glycoside hydrolase, BtGH84, we demonstrate how small molecule "fragments", identified as activators in free solution, can be covalently tethered to the protein using Michael-addition chemistry. We show how tethering generates a constitutively-activated enzyme-fragment conjugate, which displays both improved catalytic efficiency and increased susceptibility to certain inhibitor classes. Structure guided modifications of the tethered fragment demonstrate how specific interactions between the fragment and the enzyme influence the extent of activation. This work suggests that a similar approach may be used to modulate the activity of enzymes such as to improve catalytic efficiency or increase inhibitor susceptibility.

AB - Modulation of enzyme activity is a powerful means of probing cellular function and can be exploited for diverse applications. Here, we explore a method of enzyme activation where covalent tethering of a small molecule to an enzyme can increase catalytic activity (k cat/K M) up to 35-fold. Using a bacterial glycoside hydrolase, BtGH84, we demonstrate how small molecule "fragments", identified as activators in free solution, can be covalently tethered to the protein using Michael-addition chemistry. We show how tethering generates a constitutively-activated enzyme-fragment conjugate, which displays both improved catalytic efficiency and increased susceptibility to certain inhibitor classes. Structure guided modifications of the tethered fragment demonstrate how specific interactions between the fragment and the enzyme influence the extent of activation. This work suggests that a similar approach may be used to modulate the activity of enzymes such as to improve catalytic efficiency or increase inhibitor susceptibility.

UR - http://www.scopus.com/inward/record.url?scp=85032026523&partnerID=8YFLogxK

U2 - 10.1039/C7SC01966A

DO - 10.1039/C7SC01966A

M3 - Article

SN - 2041-6520

VL - 8

SP - 7772

EP - 7779

JO - Chemical Science

JF - Chemical Science

IS - 11

ER -