Fig 1: Protein·SOMAmer interactions in domain 1. A, residue Arg-16 on the IL-6 N-terminal tail hydrogen bonds to the Hoogsteen face of G29. The benzyl group of Bn-dU30 stacks against the methylene side chain of Arg-16. B, N-terminal tail of IL-6 is sandwiched between the backbone of the SOMAmer, protecting the hydrophobic modified nucleotides from solvent. Opposing strands of the SOMAmer backbone are in unusually close proximity to each other. C, Arg-24 on helix A of IL-6 forms a salt bridge to the SOMAmer backbone at G5–G6, further sealing the hydrophobic pocket from solvent. D, Tyr-31 on helix A of IL-6 stacks with Nap-dU12, which in turn stacks with Bn8 and the uridine ring of Bn-dU7. Bn7 and Bn30 have edge-to-face interactions with the stacked residues. E, Nap12 has hydrophobic interactions with the methylene side chain of Met-117 on helix C of IL-6. The naphthyl group also stacks against the uridine ring of Bn-dU8. F, Bn7 and Bn8 have edge-to-edge interactions with Phe-125 on helix C, and Bn7 interacts with the methylene side chains of Arg-24 and Lys-27.
Fig 2: A, structure of the G-quartet domain of the IL-6 SOMAmer, depicted with a C3 spacer inserted between Nap-dU12 and G29 to connect the two discontiguous sections of the domain, showing the probable structure of the G-quartet fragment. The SOMAmer is colored by B-factors from most ordered (blue) to least ordered (red) atoms. B, sequences and binding affinities of IL-6 SOMAmer (SL1025) and two G-quartet fragments (SL1028 and SL1039) connected by a C3 spacer. C, summary of systematic replacement of modified nucleotides in the G-quartet fragment (SL1028). Values shown are the ratio of the Kd values (Kdvariant/Kdparent). The Kd value for the parent fragment (SL1028) is 2.7 × 10−7 m. (N.D. = not determined; Bn-dU, 5-(N-benzylcarboxamide)-2′-deoxyuridine; iBu-dU, 5-(N-isobutylcarboxamide)-2′-deoxyuridine; FBn-dU, 5-[N-(4-fluorobenzyl)carboxamide]-2′-deoxyuridine; Pe-dU, 5-[N-(phenyl-2-ethyl)carboxamide]-2′-deoxyuridine; Pp-dU, 5-[N-(phenyl-3-propyl)carboxamide]-2′-deoxyuridine; Tyr-dU, 5-[N-(4-hydroxyphenyl-2-ethyl)carboxamide]-2′-deoxyuridine; MBn-dU, 5-[N-(3,4-methylenedioxybenzyl)carboxamide]-2′-deoxyuridine; Nap-dU, 5-[N-(1-naphthylmethyl)carboxamide]-2′-deoxyuridine; 2Nap-dU, 5-[N-(2-naphthylmethyl)carboxamide]-2′-deoxyuridine; Ne-dU, 5-[N-(1-naphthyl-2-ethyl)carboxamide]-2′-deoxyuridine; Trp-dU, 5-[N-(3-indole-2-ethyl)carboxamide]-2′-deoxyuridine; Bt-dU, 5-[N-(3-benzo[b]thiophene-2-ethyl)carboxamide]-2′-deoxyuridine; MOE-dU, 5-[N-(1-morpholino-2-ethyl)carboxamide]-2′-deoxyuridine; Bf-dU, 5-[N-(3-benzo[a]furan-2-ethyl)carboxamide]-2′-deoxyuridine; RTHF-dU, 5-[N-((R)-2-tetrahydrofurylmethyl)carboxamide]-2′-deoxyuridine; STHF-dU, 5-[N-((S)-2-tetrahydrofurylmethyl)carboxamide]-2′-deoxyuridine).
Fig 3: Bn7 only partly fills the groove on the IL-6 surface, whereas Nap12 completely fills the cavity at the protein·SOMAmer interface. Packing and shape complementarity at the IL-6·SOMAmer interface are shown in surface (IL-6) and stick representation (SOMAmer) (A) and space-filled renderings of Bn7 (B) and Nap12 (C). D, 2Fo − Fc electron density map contoured at 1.5 σ shows clear density for the aromatic stacking interactions between Tyr-31 on IL-6 and the modified nucleotides of Nap-dU12 and Bn-dU8 on the SOMAmer.
Fig 4: Detail of SOMAmer and receptor-binding sites. A, residue Phe-279 on IL-6Rα and Bn22 on the SOMAmer (domain 2) recognize the same binding site on IL-6 helices A and D (pink denotes IL-6Rα, other colors as in other figures). B, SOMAmer (domain 1) and gp130 recognize the same binding site on the IL-6 protein. Phe-169 of gp130 and Bn7, Bn8 and Nap12 of the SOMAmer interact with Tyr-31 and the methylene side chains of Leu-19 and Arg-24 on helix A of IL-6 (deep red denotes gp130). C, SOMAmer backbone between G6 and Bn7 in the IL-6·SOMAmer structure occupies the same space as Trp-142 in the IL-6·IL-6Rα·gp130 structure.
Fig 5: Protein-SOMAmer contacts in domain 2 are primarily hydrophobic. A, Bn15, Bn22, and Bn23 have hydrophobic interactions with the methylene side chains of residues on helix A and helix D on the IL-6 protein. B, Bn14 has edge-to-face interactions with Tyr-31 as well as edgewise interactions with the nonpolar side chains of Lys-27 and Arg-30. C, salt bridges between Lys-27 and Arg-30 on IL-6 and the SOMAmer backbone at A13 and Bn-dU14. D, surface rendering of IL-6 illustrates the shape complementarity of the SOMAmer·IL-6 interface.
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