*4.8. Side-Chain Conformational Analysis by Molecular Mechanics Calculations*

The conformational analysis was performed using the program Discovery Studio 2.1 (Accelrys, CA, USA) on an IBM x3550M2 workstation (CPU: Dual Xeon E5530 2.4 GHz with Quad cores; RAM: 48 G) running the operating system CentOS 5.3. The models were created based on the solution structure of an analogous peptide of the parent YKL peptide [46] with various combinations of potential low-energy side-chain dihedrals. For each side-chain dihedral angle (χ) involving sp<sup>3</sup> carbons, three possible low-energy staggered conformations were considered: gauche− (60◦ , g−), trans (180◦ , t), and gauche+ (300◦ , g+) [47,48]. For the dihedral angle involving the sp<sup>2</sup> carboxylate carbon of Aad, six conformations were considered: 0◦ , 30◦ , 60◦ , 90◦ , 120◦ , and 150◦ . For each peptide (HPTAadDab and HPTDabAad), 1458 conformations were evaluated. Each conformation was minimized using the CFF forcefield. The nonbond radius of 99 Å, nonbond higher cutoff distance of 98 Å and nonbond lower cutoff distance of 97 Å were employed to perform the calculations with effectively no cutoffs. Distance dependent dielectric constant of 2 was used as the implicit solvent model. Minimization was performed by steepest descent and conjugate gradient protocols until convergence (converging slope was set to 0.1 kcal/(mol × Å). After

minimization, each conformation was reexamined to remove duplicating conformations because minimization with different starting conformations occasionally resulted in the same final conformation. When the same conformation was obtained more than once, only the lowest energy conformation was considered in further analyses. The probability of conformation i at 298 K (p<sup>i</sup> ) was calculated based on Boltzmann distribution using Equation (8), in which ε<sup>i</sup> is the energy of conformation i, k<sup>B</sup> is the Boltzmann constant, and T is the temperature (298 K). The entropic contribution to the folded form at 298 K was calculated using Equation (9).

$$\mathbf{p}\_{\mathbf{i}} = \frac{\mathbf{e}^{\frac{-\epsilon\_{\mathbf{i}}}{\mathbf{k}\_{\mathbf{B}^{\mathsf{T}}}}}}{\sum\_{\mathbf{j}} \mathbf{e}^{\frac{-\epsilon\_{\mathbf{j}}}{\mathbf{k}\_{\mathbf{B}^{\mathsf{T}}}}}} \tag{8}$$

$$-\text{TS} = -\text{T} \cdot (-\text{k}\_{\text{B}}) \sum\_{\text{i}} \mathbf{p}\_{\text{i}} \cdot \ln(\mathbf{p}\_{\text{i}}) \tag{9}$$
