Carry eclipse fluorescence spectrophotometer

The secondary and tertiary structure profile of the purified and refolded mature version recombinant serratiopeptidase was analyzed through circular dichroism and fluorescence spectroscopy. In brief, one micromolar recombinantly prepared serratiopeptidase was scanned in Far-UV circular dichroism spectra, i.e. 200–250 nm in 1 mm path length cell using J-810 spectropolarimeter (Jasco, UK) flushed with nitrogen gas at 25 °C. Samples were scanned at a rate of 50 nm/min with a step size of 1 nm. Spectra were averaged over three scans and corrected for background by subtracting the scans of the buffer without protein.
Three independent intrinsic tryptophan fluorescence emission spectra of one micromolar protein were collected between 300 and 450 nm after excitation at 280 nm at 25 °C using carry eclipse fluorescence spectrophotometer (Agilent technologies USA) and averaged out. The circular dichroism profile and intrinsic fluorescence spectra in the same range given by one micromolar commercial version serratiopeptidase (Systopic Laboratories, India) were taken as a control for comparison.

High-resolution mass spectrometry (HRMS) involved the use of an electrospray ionization mass spectrometer (MicroTOF, Bruker Daltonics, Billerica, USA). ¹H NMR and ¹³C NMR spectra were recorded on a JEOL JNM-ECZR (Jeol, USA) at 500 MHz and 125 MHz, respectively. The UV-visible spectra were obtained on a Varian Cary 50 UV-vis spectrophotometer (Varian, USA). The emission spectra were obtained from a Carry Eclipse fluorescence Spectrophotometer (Agilent Technologies). IR spectra results were acquired from the ALPHA II Compact Fourier-transform infrared (FTIR) Spectrometer (Bruker, Billerica, USA). The pH values of the experimental solution were measured from an Ohaus pH meter, and the UV (λ_ex = 312 nm) and blacklight (λ_ex = 365 nm) sources from standard transilluminator TCP-20.LM V1 UV 365/312 nm (Vilber Lourmat, Collégien, France) were used for naked eye observations in the sensing experiments. The optical and morphological characterization of AuNPs were obtained from the Dynamic Light Scattering (DLS) of a Zetasizer Nano ZSP (Malvern, UK) with a 1.00 cm quartz cell at room temperature and Transmission Electron Microscopy (TEM) of a JEM-1400 electron microscope under an accelerating voltage of 120 kV (Jeol, USA).