Nmr spectrometer

Infrared spectra were recorded on a
PerkinElmer spectrum 100 spectrophotometer. High-resolution ESI mass
spectra were recorded on an Agilent 6220 Accurate Mass TOF LC-MS spectrometer.
Melting points were determined using a Reichert apparatus and are
uncorrected. Nuclear magnetic resonance (NMR) spectra were recorded
on a 300, 400, or 600 MHz Bruker NMR spectrometers in CDCl₃ at 298 K (unless stated otherwise). All chemical shift values are
reported in parts per million (ppm) with coupling constant (J) values reported in Hz. All spectra were referenced to
TMS for ¹H NMR and the CDCl₃ solvent peak for ¹³C{¹H} NMR. The anhydrous solvents were purchased
from commercial sources and used as received. TLC tests were run on
TLC Alugram Sil G plates and visualized under UV light at 254 nm.
Chromatography: Separations were carried out on silica gel. The general
procedures and characterization for the substrates 1a–o are included in the Supporting Information.

We determined initial rates of deamination reaction by using ¹H NMR spectra. A 9nt ssDNA substrate (IDT), 5ʹ-AATCCCAAA, was used to determine the reaction rate. NMR spectra were acquired at 25 °C on Bruker NMR spectrometers operating at ¹H Larmor frequencies of 600 and 800 MHz. NMR samples contained 5% deuterium oxide with 200 nM protein, 200 µM ssDNA substrate, 100 mM NaCl, 0.002% Tween 20, 1 mM DTT, 10 µM ZnCl₂, and also included 50 mM sodium phosphate adjusted to pH 7.5. The concentration of deamination product (5ʹ-AATCCdeoxy-UAAA) was determined from integration of the H5 uracil proton peaks at 5.60 ppm. A series of ¹H spectra were measured and the product concentrations as a function of the reaction times were used to determine the initial rate via linear regression.

The chemicals were purchased from Sigma-Aldrich and used without purification. A Nicolet iS10 spectrometer from Thermo Scientific was used to record IR spectra (KBr). The ¹H NMR and ¹³C NMR spectra were recorded by JEOL's (500 MHz) and Bruker NMR spectrometers (400 MHz) in DMSO-d₆ and CDCl₃. Mass spectra was measured through Thermo Scientific GC/MS model ISQ. The Perkin-Elmer 2400 analyzer was used to obtain the elemental analysis of C, H, and N.

Unless otherwise stated, all chemicals were purchased from commercial suppliers and used without further purification. The solvents were purified by conventional methods before used. All analytes were obtained by traditional methods. 4-Chloro-2, 1, 3-benzoxadiazole was purchased from TCI chemical. Silica gel (200–300 mesh) used for flash column chromatography was purchased from Qingdao Haiyang Chemical Co., Ltd. ¹HNMR and ¹³CNMR spectra were determined by 400 MHz and 100 MHz using Bruker NMR spectrometers. Chemical shifts (δ) were expressed as parts per million (ppm, in CDCl₃ or DMSO, with TMS as the internal standard). Meanwhile, high-resolution mass spectrometry was achieved with ESI-TOF and FTMS-ESI instrument. Fluorescence measurements were performed on an Agilent Technologies CARY Eclipse fluorescence spectrophotometer, and absorption spectra were measured on a PerkinElmer Lambda 35 UV–vis spectrophotometer. The pH values of sample solutions were measured with a precise pH-meter pHS-3C. Fluorescence quantum yield was achieved from a C11347-11 Absolute PL Quantum Yield Spectrometer. MTT assays were conducted on the Varioskan LUX Multimode Microplate Reader. The instrument used for imaging living cells and tissues of mice was an Olympus FV 1000 confocal microscopy purchased from Olympus.