Multipoint Immobilization at Inert Center of Papain on Homo-Functional Diazo-Activated Silica Support: A Way of Restoring ?Above Room-Temperature? Bio-Catalytic Sustainability

Although enzymes play a significant role in industrial applications, their potential usage at high-level efficiency, particularly above room temperature, has not yet been fully harnessed. It brings above room-temperature catalytic sustainability of an immobilized (imm.) bio-catalyst as a long pending issue to improve enzyme stability, activity, specificity, or selectivity, particularly the enantio-selectivity over the native-enzymes. At this juncture, in a robust methodology, a heterogeneous solid phase bio-catalyst, {Si(OSi)4(H2O)1.03}n= 3 2 8{OSi(CH3)2-NH-C6H4-N= N}4{papain}(H2O)251, has efficiently been prepared by immobilizing papain on homo-functionalized SG (silica-gel) via multipoint covalent attachment. The bio-catalyst is easy to be recovered and reused multiple times. The homo-functional -N=N+, which appears on the SG-surface, makes the multipoint diazo-links with the inert center of the tyrosine-moiety to couple the enzyme where all the amino, thiol, phenol, and so forth, groups of the protein, including those that belong to the active-site, remain intact. The immobilized enzyme (13.9 smol g-1) swims in pore-water within the pore-channel, remains stable up to 70 +/- 5 degrees C, and exhibits wider temperature adaptability in performing its hydrolyzing activities. The relative activity, 78 +/- 2% at 27 degrees C, remains quantitative for 60 days and can be reused for 60 cycles with 53% activity at room-temperature. The thermal (relative activity: 87%; incubated at 70 +/- 5 degrees C for 24 h) and mechanical (relative activity: 92%; incubated at 2500 rpm for 2 h at 27 degrees C) stability was outstanding.

Automated summary

Enzymes have great potential in industrial applications, but utilizing them at high efficiency levels, especially above room temperature, has not been fully realized. The immobilization of papain onto homo-functionalized silica gel via multipoint covalent attachment has successfully addressed this issue, resulting in a stable and reusable enzyme catalyst with improved activity and adaptability to higher temperatures.