. Author manuscript; available in PMC: 2009 Nov 5.

Published in final edited form as: J Am Chem Soc. 2008 Oct 9;130(44):14675–14683. doi: 10.1021/ja804656h

Table 1.

Effects of Termini Changes on the NMR Structuring Diagnostics of –W-loop-W- Peptides.

`#`	`Sequence`	`Tm (°C)^a`	`f_F^b`	`CSD (ppm)`
		`228 nm CD melt`	`<frCSD>`	`S-2 W Hε3^c`	`S+2 W Hε3^c`	`S+4 HN_E`
`1`	`Ac-WNPATGKW-NH₂`		~0.45 (~0.48 ^CD)^c	−0.873	−0.179	n. a.
`2`	`KAVW-INGK-WTVE`	~65	~0.83	−1.922^d	−0.280	+0.55
`3`	`Ac-W-INGK-W-NH₂`	~5	≤0.45 (~.42 ^CD)^c	−0.378	−0.738	n. a.
`4`	`Ac-W-IpGK-W-NH₂`	≥ 45	.80 ± 0.05	−0.26	−1.64	n. a.
`5`	`Ac-TW-INGK-W-NH₂`		≤0.30 (~.1 ^CD)	−0.570	−0.384	n. a.
`6`	`Ac-W-INGK-WT-NH₂`	~41	0.76 ± 0.04	−0.380	−1.680	−2.7^c
`7`	`Ac-W-INGK-WTG-NH₂`	~38	0.77 ± 0.04	−0.379	−1.556	−2.71
`8`	`W-INGK-WTG`		0.15 ± 0.10	−0.052	−0.108	−1.11
`9`	`W-INGK-WTG-NH₂`		0.09 ± 0.09	−0.017	−0.067	−1.17
`10`	`Ac-WTG-NH₂`					−1.26
`11`	`Ac-W-IpGK-WTG-NH₂`	≥ 65	0.94 ± 0.03	−0.25	−1.87	−3.06
`12`	`Ac-W-IpnK-WTG-NH₂`	≥ 68	< 0.97 >	−0.31	−1.93	−3.20 ^e

These Tm estimates assume that 100% folded [θ]₂₂₈ value is temperature independent and, with the exception of entries 2, 4 and 12, are derived from Figure S3 of reference ¹⁴. Variation in the 100%-folded value with sequence changes^b,c, adds additional uncertainty to these estimates.

The reported <frCSD> values are the average (and standard error) of the 280K f_F values derived from strand backbone H_N and Hα sites displaying moderate to large CSDs (W1 Hα&H_N, I2 Hα&H_N, K5 Hα, W6 Hα&H_N) assuming that Ac-WIPGKWTG-NH₂ represents zero folding and that the best folded species (entry #12, Ac-WIpnKWTG-NH₂) represents f_F = 0.97. CD measures of f_F were based on the magnitude of the 228 nm maximum of the W/W exciton couplet; the 100%-folded value calculated for entry #2 is, in Molar (not residue-Molar) terms, +446,000°¹⁴. The 100% [θ]₂₂₈ value for entry #12 is +303,000°; an increase (to +418,000° at 278K) was observed for the optimized construct (Pr-WIpGIWTGPS) reported herein.

The largest [θ]₂₂₈ value observed for prior-WNPATGKW- species is +434,000°,^10c the CD folding estimate in entry #2 is based on this value. Other analogs with this W-loop-W motif have 100%-folded [θ]₂₂₈ values as small as +380,000°. The maximum 100% value observed for W-(S/I)-(N/p)-GK-W species was +490,000°.^10c Recent studies¹⁴, have documented both WIpGKW- and WINGKW-containing species in which the 100%-folded values are ≤ +300,000° (calibrated based on NMR measures of folding); it appears that magnitude of the exciton couplet can vary significantly.

CSDs for the shielded S±2 Trp Hε3 as large as −2.3 ppm have been observed for longer peptide hairpins incorporating the …W-turn-W … sequence (“turn”= NPATGK or INGK).^10c,¹⁴ Variations in the preferred W/W interaction geometry can give rise to slightly different 100% folded values.¹⁴

The largest upfield shift observed for the Gly H_N in prior WTG-NH₂ terminated peptides was 3.42 ppm.¹⁴ In the present study, one construct shows a 3.91 ppm upfield shift under the same conditions (20 vol-% glycol at 270K.)