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In mechanism-based inhibition, catalytic processing of an inhibitor generates the species responsible for sustained inhibition. The covalent bond can be between inhibitor and enzyme cofactor rather than to an amino acid side chain; persistence must be described on the relevant timescale.

finasteride is reduced at C1, leaving a C2 enolate that attacks nicotinamide C4 to make the covalent C–C adduct. In the inhibitor's unsaturated lactam, the reactive chain is $\mathrm{O}=C3-C2=C1$, with N at steroid position 4. Activate the C3 carbonyl by hydrogen bonding; draw the reduced cofactor's C4–H bond toward inhibitor C1, the C1=C2 pi pair toward C2–C3, and the C3=O pi pair toward oxygen. This gives NADP+ next to the C2-centered resonance form of the inhibitor enolate.

Instead of completing reduction by protonating C2, move the C2 lone pair toward the electrophilic C4 of the oxidized nicotinamide ring. Shift the pyridinium pi system toward its positively charged N, and reform the inhibitor carbonyl if using the oxygen-centered enolate representation. The result is the C2(inhibitor)–C4(nicotinamide) bond and a neutral dihydronicotinamide ring. Numbering the two molecules separately prevents confusing cofactor C4 with the inhibitor's N4. The lactam N–H remains present and can help position the inhibitor through hydrogen bonding; it does not form a covalent bond to the protein.

This is mechanism-based inhibition: enzyme chemistry generates the species that traps its own oxidized cofactor. Isolation of a cofactor–inhibitor adduct after protein denaturation, rather than persistent attachment to a denatured protein, distinguishes it from covalent modification of an amino acid side chain. The original mechanistic study combined radiolabel recovery, purification and spectroscopy to establish the adduct; its slow breakdown also cautions against treating “irreversible” as an absolute statement for every timescale.^[ref]

Write concentrations as $E,S,X,Y,E_i,P$ in this calculation. The law of mass action assigns the forward bimolecular rate $k_{1}ES$ and the first-order rates $k_{-1}X,k_{2}X,k_{3}Y,k_{4}Y$. Each species gains the rates of reactions producing it and loses those consuming it:Adding the equations for all enzyme-bearing species cancels every reaction rate, so $\boxed{E+X+Y+E_i=e_0}$. This is conservation of the total enzyme-derived material, including the inactivated form. It does not assert conservation of active enzyme: mechanism-based inhibition transfers some of that conserved material irreversibly into $E_i$.

A quasi-steady-state approximation for enzyme-substrate complexes can remain useful when total enzyme and initial substrate concentrations are comparable. The relevant parameter can involve the sum $s_0+K_m$, rather than $s_0$ alone. In a mechanism-based inhibition model with $s_0/K_m=O(\epsilon )$, bounded intermediate decay rates and an inactivation rate of order $\epsilon$, the fast complexes adjust in a layer of scaled width $O(\epsilon )$. On bounded slow-time intervals the main complex follows the substrate, the later complex and accumulated inactivated enzyme remain $O(\epsilon )$, and the substrate has leading exponential depletion. An outer algebraic complex concentration does not satisfy the unbound initial condition inside that initial layer.