Generating monomeric 5-coordinated microperoxidase-11 using carboxylic acid functionalized silver nanoparticles: A surface-enhanced resonance Raman scattering analysis

Microperoxidase-11 (MP-11), a heme undecapeptide obtained by proteolytic digestion of cytochrome c, resembles peroxidase enzyme when its heme center is 5-coordinated with a vacant sixth coordination site. However, MP-11 always tends to aggregate in both solution and on surface and eventually forms the 6-coordinated heme. Thus, the present study investigates the immobilization strategy of MP-11 on nanoparticle surface in order to generate monomeric 5-coordinated MP-11 and make them as an efficient biocatalyst. The powerful surface-enhanced resonance Raman scattering (SERRS) technique is being employed to attain the detailed structural information of the catalytic site i.e., the heme center. The localized surface plasmon resonance (LSPR) tuned and 6-mercaptohexanoic acid (MHA) functionalized silver nanoparticles (Ag@MHA NPs) are used as Raman signal amplifier. The outcome of the SERRS study unambiguously portrays the existence of monomeric 5-coordinated MP-11 on Ag@MHA NPs surface. Here, Ag@MHA NPs plays a dual role of providing a platform to create monomeric 5-coordinated MP-11 and to load large number of MP-11 due to its high surface to volume ratio. Further, the electrostatic interaction between Ag@MHA NPs and MP-11 leads to instantaneous SERRS signal enhancement with a Raman enhancement factor (EFSERS) of 2.36×10(6). Langmuir adsorption isotherm has been employed for the adsorption of MP-11 on Ag@MHA NPs surface, which provides the real surface coverage (ΓS(*)) and equilibrium constant (K) value of 1.54nm and 5×10(11)M(-1). Furthermore, the peroxidase activity of MP-11 has been demonstrated through electrocatalytic oxygen reduction reaction.