Functional determinants of human enteric α-defensin HD5: Crucial role for hydrophobicity at dimer interface

Mohsen Rajabi, Bryan Ericksen, Xueji Wu, Erik De Leeuw, Le Zhao, Marzena Pazgier*, Wuyuan Lu

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

63 Scopus citations


Human α-defensins are cationic peptides that self-associate into dimers and higher-order oligomers. They bind protein toxins, such as anthrax lethal factor (LF), and kill bacteria, including Escherichia coli and Staphylococcus aureus, among other functions. There are six members of the human α-defensin family: four human neutrophil peptides, including HNP1, and two enteric human defensins, including HD5. We subjected HD5 to comprehensive alanine scanning mutagenesis. We then assayed LF binding by surface plasmon resonance, LF activity by enzyme kinetic inhibition, and antibacterial activity by the virtual colony count assay. Most mutations could be tolerated, resulting in activity comparable with that of wild type HD5. However, the L29A mutation decimated LF binding and bactericidal activity against Escherichia coli and Staphylococcus aureus. A series of unnatural aliphatic and aromatic substitutions at position 29, including aminobutyric acid (Abu) and norleucine (Nle) correlated hydrophobicity with HD5 function. The crystal structure of L29Abu-HD5 depicted decreased hydrophobic contacts at the dimer interface, whereas the Nle-29-HD5 crystal structure depicted a novel mode of dimerization with parallel β strands. The effect of mutating Leu 29 is similar to that of a C-terminal hydrophobic residue of HNP1, Trp26. In addition, in order to further clarify the role of dimerization in HD5 function, an obligate monomer was generated by N-methylation of the Glu21 residue, decreasing LF binding and antibacterial activity against S. aureus. These results further characterize the dimer interface of the α-defensins, revealing a crucial role of hydrophobicity-mediated dimerization.

Original languageEnglish
Pages (from-to)21615-21627
Number of pages13
JournalJournal of Biological Chemistry
Issue number26
StatePublished - 22 Jun 2012
Externally publishedYes


Dive into the research topics of 'Functional determinants of human enteric α-defensin HD5: Crucial role for hydrophobicity at dimer interface'. Together they form a unique fingerprint.

Cite this