. 2018 Jul 17;2(3):036102. doi: 10.1063/1.5022841

TABLE I.

Characterization data for PEG conjugates.

		¹H NMR			K_d (μM)^e
	Yield (wt.%)^a	Pept./ME (mol %)^b	$\bar{M_{n}}$ (g/mol)^c	MALDI-ToF (m/z)^d	ITC	FA
PEG-VS	72	…	2089	[M+Na]⁺ 2126	…	…
PEG-VS	72	…	2089	[M+K]⁺ 2110	…	…
PEG-pep1	66	80/20	2851	[M]⁺ 2686	103 ± 5 Pep1: 44 ± 9	135 ± 8 Pep1: 40 ± 1
PEG-pep2	60	86/14	2891	[M]⁺ 2770	Not detected Pep2: 351 ± 10	1.8± 0.6 × 10³ Pep2: 335 ± 6
PEG-pep3	90	70/30	2662	[M+Na]⁺ 2727[M+H]⁺ 2705	Not measurable Pep3: not detected	Not measurable Pep3: not detected

^{^a}

Calculated as the weight of recovered material/theoretical weight of the reaction product.

^{^b}

Proton nuclear magnetic resonance (¹H-NMR) spectra, chemical structures, and proton numbering are reported in supplementary material, Fig. 7SI. The ratio between peptide- and ME-terminated PEG is calculated from the ratio of the resonance of a PEG-ME methylene at 2.84 ppm (protons A, in α to the thioether and β to the terminal primary alcohol) and the PEG-pep proline main chain protons either at 4.54 (H19) and 4.47 (H11) ppm (for PEG-pep1), at 4.48 (H11) ppm (for PEG-pep2) or at 4.57–4.45 (H11, H17) ppm (for PEG-pep3); the vinyl sulfone peak has completely disappeared.

^{^c}

PEG-VS samples: $\bar{M_{n}}$ calculated using the number of repeating units calculated by averaging the ratio of the ¹H NMR resonance of the PEG-VS terminal groups (methylene and ethylene protons at 3.25 ppm and 6.1/6.4 ppm) and those in the main chain at 3.5–3.8 ppm. PEG-peptide samples: $\bar{M_{n}}$ calculated from the ratio of the ¹H NMR resonances of terminal groups [obtained averaging the PEG-ME methylene (protons A) or PEG-peptides proline protons (H11, H17, H19)], and those of PEG main chain.

^{^d}

Most intense peaks in MALDI-ToF (Matrix-Assisted Laser Desorption/Ionization-Time of Flight) spectra; all other peaks in the distribution are spaced by an additive 44 mass atomic units (see also supplementary material, Fig. 6SI).

^{^e}

K_d values obtained through fluorescence anisotropy (FA) and isothermal titration calorimetry (ITC) as described in the Methods section and in supplementary material, Sec. 1SI. The values obtained for non-conjugated peptides are also reported; note that Pep3 showing no binding, no competition assay is possible for PEG-pep3. n = 3 independent samples for both techniques.