Influence of Structural Features of Peptides on Their Affinity to Cotton Linters Paper

As an alternative to chemical functionalization of cellulose, fusion constructs with carbohydrate-binding modules (CBMs) can be used for the noncovalent immobilization of various compounds and functionalities on cellulose. Smaller cellulose-binding peptides might be used as an alternative, as they are easy to modify and can be produced completely synthetically. To investigate which structural features of peptides promote binding to cotton linters paper, we have established a label-free assay to assess paper affinity. Even though tyrosine residues are essential for the binding of CBMs, we found that the Y/A exchange in peptides did not lead to a reduction in the affinity. This confirms previous assumptions that aromatic structures that are needed to orient CBMs during cellulose binding are less important for smaller, more flexible peptides. We also show that the aromatic fluorophore 5(6)-carboxytetramethylrhodamine (TAMRA), which is sometimes used for peptide labeling, leads to an increase in affinity due to an avidity effect. In addition, we observed that peptides with a <i>C</i>-terminal carboxylate group or carboxylate side chains have a lower affinity, and peptides with positively charged amino groups have a higher affinity than the corresponding uncharged peptides. We attributed this to electrostatic interactions with carboxylate groups on the paper. While most peptides tested in this study bound with <i>K</i> _D values in the midmicromolar range, the combination of a <i>C</i>-terminal amide and an <i>N</i>-terminal TAMRA modification yielded a peptide with affinity to cotton linters paper in the low micromolar range. The findings presented in this work confirm and expand findings from previous work and reveal limitations and features that point the way toward high-affinity peptides for the functionalization of cellulose.