Collagen is a key structural protein in the extracellular matrix of many tissues. It provides biological tissues with tensile mechanical strength and is enzymatically cleaved by a class of matrix metalloproteinases known as collagenases. Collagen enzymatic kinetics has been well characterized in solubilized, gel, and reconstituted forms. However, limited information exists on enzyme degradation of structurally intact collagen fibers and, more importantly, on the effect of mechanical deformation on collagen cleavage. We studied the degradation of native rat tail tendon fibers by collagenase after the fibers were mechanically elongated to strains of . After the fibers were elongated and the stress was allowed to relax, the fiber was immersed in Clostridium histolyticum collagenase and the decrease in stress was monitored as a means of calculating the rate of enzyme cleavage of the fiber. An enzyme mechanokinetic (EMK) relaxation function in was calculated from the linear stress-time response during fiber cleavage, where corresponds to the zero order Michaelis–Menten enzyme-substrate kinetic response. The EMK relaxation function was found to decrease with applied strain at a rate of per percent strain, with complete inhibition of collagen cleavage predicted to occur at a strain of . However, comparison of the EMK response ( versus ) to collagen’s stress-strain response ( versus ) suggested the possibility of three different EMK responses: (1) constant within the toe region , (2) a rapid decrease in the transition of the toe-to-heel region followed by (3) a constant value throughout the heel and linear regions. This observation suggests that the mechanism for the strain-dependent inhibition of enzyme cleavage of the collagen triple helix may be by a conformational change in the triple helix since the decrease in appeared concomitant with stretching of the collagen molecule.