Table 1.
Constructs analysed in this study
| H2′ name | H2′ sequence | H name | H sequence | ΔGapp (kcal/mol) |
|---|---|---|---|---|
|
H2′ control experiments | ||||
| Wild type |
…TGASVFPVLAIVLIV… |
3L/16A |
AAAALAAAALAAAALAAA |
0.1; ND |
| CCH2 |
…LAIVLIVSPSAQA+AY… |
3L/16A |
AAAALAAAALAAAALAAA |
−0.2; ND |
| pPL |
pPL signal peptide |
3L/16A |
AAAALAAAALAAAALAAA |
0.1; ND |
| 19L |
LLLLLLLLLLLLLLLLLLL |
3L/16A |
AAAALAAAALAAAALAAA |
0.1; ND |
| 1D/18L |
LLLLLLLLLDLLLLLLLLL |
3L/16A |
AAAALAAAALAAAALAAA |
−0.3; ND |
| 1N/18L |
LLLLLLLLLNLLLLLLLLL |
3L/16A |
AAAALAAAALAAAALAAA |
−0.2; ND |
| 1D/18L |
LLLLLLLDLLLLLLLLLLL |
3L/16A |
AAAALAAAALAAAALAAA |
−0.2; ND |
| 1N/18L |
LLLLLLLNLLLLLLLLLLL |
3L/16A |
AAAALAAAALAAAALAAA |
−0.0; ND |
| 2D/17L |
LLLLLLLDLLLDLLLLLLL |
3L/16A |
AAAALAAAALAAAALAAA |
−0.0; ND |
| 2N/17L |
LLLLLLLNLLLNLLLLLLL |
3L/16A |
AAAALAAAALAAAALAAA |
−0.0; ND |
|
Single Asp and Asn experiments | ||||
| 19L |
LLLLLLLLLLLLLLLLLLL |
1N/5L/13A (N) |
AAAAAAALLNLLLAAAAA |
0.3; 0.8 |
| 1N/18L |
LLLLLLLLLNLLLLLLLLL |
1N/5L/13A (N) |
AAAAAAALLNLLLAAAAA |
0.2; 0.8 |
| CCH2 |
…LAIVLIVSPSAQA+AY… |
1N/5L/13A (N) |
AAAAAAALLNLLLAAAAA |
0.0, ND |
| 19L |
LLLLLLLLLLLLLLLLLLL |
1D/7L/11A (D) |
AAAAAALLLDLLLLAAAA |
0.1; 0.4 |
| 1D/18L |
LLLLLLLLLDLLLLLLLLL |
1D/7L/11A (D) |
AAAAAALLLDLLLLAAAA |
−0.1; 0.3 |
|
Double Asp and Asn experiments | ||||
| 19L |
LLLLLLLLLLLLLLLLLLL |
2N/8L/9A (N5Nl) |
AAAALLNLLLLLNLAAAA |
−0.1; ND |
| 2N/17L |
LLLLLLLNLLLNLLLLLLL |
2N/8L/9A (N5Nl) |
AAAALLNLLLLLNLAAAA |
−0.3; ND |
| 19L |
LLLLLLLLLLLLLLLLLLL |
2N/9L/8A (N5Ns) |
AAAALLNLLLLLNLLAAAA |
ND; −0.5 |
| 2N/17L |
LLLLLLLNLLLNLLLLLLL |
2N/9L/8A (N5Ns) |
AAAALLNLLLLLNLLAAAA |
ND; −0.7 |
| 19L |
LLLLLLLLLLLLLLLLLLL |
2N/9L/8A (N3N) |
AAAALLLNLLLNLLLAAAA |
0.3; 0.6 |
| 2N/17L |
LLLLLLLNLLLNLLLLLLL |
2N/9L/8A (N3N) |
AAAALLLNLLLNLLLAAAA |
0.1; 0.0 |
| 19L |
LLLLLLLLLLLLLLLLLLL |
2N/9L/8A (N1N) |
AAAALLLLNLNLLLLAAAA |
−0.3; −0.2 |
| 2N/17L |
LLLLLLLNLLLNLLLLLLL |
2N/9L/8A (N1N) |
AAAALLLLNLNLLLLAAAA |
−0.5; −0.6 |
| 19L |
LLLLLLLLLLLLLLLLLLL |
2D/11L/6A (D5D) |
AAALLLDLLLLLDLLLAAA |
−0.3; −0.1 |
| 2D/17L |
LLLLLLLDLLLDLLLLLLL |
2D/11L/6A (D5D) |
AAALLLDLLLLLDLLLAAA |
−0.4; −0.2 |
| 19L |
LLLLLLLLLLLLLLLLLLL |
2D/12L/5A (D3D) |
AAALLLLDLLLDLLLLLAA |
0.0; 0.0 |
| 2D/17L |
LLLLLLLDLLLDLLLLLLL |
2D/12L/5A (D3D) |
AAALLLLDLLLDLLLLLAA |
−0.5; −0.9 |
| 19L |
LLLLLLLLLLLLLLLLLLL |
2D/12L/5A (D1Dl) |
AAALLLLLDLDLLLLLLAA |
−1.0; ND |
| 2D/17L |
LLLLLLLDLLLDLLLLLLL |
2D/12L/5A (D1Dl) |
AAALLLLLDLDLLLLLLAA |
−1.1; ND |
| 19L |
LLLLLLLLLLLLLLLLLLL |
2D/11L/6A (D1Ds) |
AAALLLLLDLDLLLLLAAA |
ND; 0.2 |
| 2D/17L | LLLLLLLDLLLDLLLLLLL | 2D/11L/6A (D1Ds) | AAALLLLLDLDLLLLLAAA | ND; −0.1 |
The H2′ and H segments are listed for each construct (abbreviated H segment names in parentheses refer to Figure 2). All H2′ segments (except wild type, CCH2 and pPL) and all H segments have GGPG…GPGG flanks. Two ΔGapp values are given: the first refers to constructs with a 150-residue-long loop between H2′ and H segments and the second to constructs with a 55-residue loop (ND, not done). Pairs of constructs used to calculate the ΔΔGapp values shown in Figure 2 are indicated. Note that the composition of the H segments are in some cases different for the long- and short-loop constructs (indicated by subscripts l and s in the abbreviated H segment names, respectively), necessitated by the requirement that ΔGapp≈0 kcal/mol. The standard deviation in repeat ΔGapp measurements is σ≈±0.1 kcal/mol (Hessa et al, 2005a). A, Ala, alanine; D, Asp, aspartic acid; L, Leu, leucine; N, Asn, asparagine.