A rational strategy is presented here to enhance enzyme stability by gaining control over the noncovalent interactions at the enzyme–graphene interface (EGI). The charge (n) on a model enzyme, glucose oxidase (GOx), was systematically varied from −67 to +78 via chemical modification of its COOH groups with polyamines, and chemically modified GOx­(n) has been adsorbed onto graphene oxide (GO). Control of the net charge on GOx­(n) provided an excellent handle to engineer the EGI to enhance the stability of the GOx­(n)/GO biocatalyst while retaining full activity. Enzyme loading (w/w, GOx­(n) to GO) increased with increased n, and a maximum enzyme loading of 420% (w/w) was noted when n = 0; a further increase in n decreased loading. Enzymatic activities of the GOx­(n)/GO hybrids increased steadily with n, and the maximum specific activity was about 1.6 times greater than that of GOx. There has been a good correlation between the retention of enzyme secondary structure and the corresponding enzymatic activities. These indicated an unprecedented increase in kinetic stability as a function of n, measured at 40 °C, with a maximum half-life of 38 days (n = 0, +35), which is about 150 times higher than that of GOx, under the same conditions. The kinetic barrier to denaturation increased steadily from 58.7 kcal/mol for GOx to 294 kcal mol–1 for GOx­(+35)/GO and then decreased slightly with increased n. Chemical denaturation studies showed that the improved stability is not due to changes in thermodynamic stability and that the increased kinetic stability is due to an increased barrier to denaturation. Adsorption onto GO played a key role in enhancing enzyme stability, and without GO, the stability enhancements were only marginal. Thus, engineering the EGI provided exciting opportunities to generate highly stable biocatalysts that can be stored under ambient conditions, while retaining high activities, with very long lifetimes, much needed for practical applications.