A new bioproduction route for a novel antimicrobial peptide.
Beta defensins are antimicrobial peptides (AMPs) with a broad spectrum antimicrobial behavior against pathogens while having minimal tendency to incur pathogen resistance. Human β-defensin 28 (hBD28) is a strongly cationic AMP and hence hypothesized to be highly effective in permeabilizing negatively-charged pathogen membranes. However, the scarcity of hBD28 in vivo has impeded detailed structure and antimicrobial studies of hBD28. Chemical synthesis of hBD28 rendered extremely poor yields due to inefficient cysteine oxidation. In this study, a rapid and scalable production route to produce bioactive hBD28 in Escherichia coli (E. coli) is reported. The design of a dual fusion tag expression construct was pivotal in enhancing soluble expression and easing purification of hBD28. The final hBD28 (purity >95%) displayed significant antimicrobial activity against E. coli K12 and showed dose-dependent killing kinetics. Circular dichroism spectroscopy confirmed the presence of both β-sheet and α-helix conformations in the secondary structure of hBD28.
School of Chemical and Biomedical Engineering, Nanyang Technological University, Singapore.
Derrick Kok Sing
DK
Tay
Gobinath
G
Rajagopalan
Xiang
X
Li
Yu
Y
Chen
Linda H L
LH
Lua
Susanna Su Jan
SS
Leong
D000890:Anti-Infective Agents / Q000737:chemistry* / Q000302:isolation & purification*; D001709:Biotechnology / Q000379:methods*; D002942:Circular Dichroism; D048168:Escherichia coli K12 / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D023083:beta-Defensins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification
D000890:Anti-Infective Agents; D011993:Recombinant Fusion Proteins; D023083:beta-Defensins