Affinity comparison of p3 and p8 peptide displaying bacteriophages using surface plasmon resonance.
Ever increasing demands in sensitivity and specificity of biosensors have recently established a trend toward the use of multivalent bioreceptors. This trend has also been introduced in the field of bacteriophage affinity peptides, where the entire phage is used as a receptor rather than the individual peptides. Although this approach is gaining in popularity due to the numerous advantages, binding kinetics of complete phage particles have never been studied in detail, notwithstanding being essential for the efficient design of such applications. In this paper we used an in house developed fiber-optic surface plasmon resonance (FO-SPR) biosensor to study the affinity and binding kinetics of phages, displaying peptide libraries. By using either peptide expression on the p3 or on the p8 coat proteins, a corresponding density of 5 up to more than 2000 peptides on a single virus particle was obtained. Binding parameters of 26 different filamentous phages, displaying peptides selective for enhanced Green Fluorescent Protein (eGFP), were characterized. This study revealed a broad affinity range of phages for the target eGFP, indicating their potential to be used for applications with different requirements in binding kinetics. Moreover, detailed analysis of koff and kon values of several selected p3 and p8 phages, using the FO-SPR biosensor, clearly showed the correlation between the binding parameters and the density at which eGFP-peptides are being expressed. Consequently, although p3 and p8-based phages both revealed exceptionally high affinities for eGFP, two p8 phages were found to have the highest affinity with dissociation constants (Kd) in the femtomolar range.
BIOSYST-MeBioS, KU Leuven , Willem De Croylaan 42, P.O. Box 2428, B-3001 Leuven, Belgium.
Karel
K
Knez
Wim
W
Noppe
Nick
N
Geukens
Kris P F
KP
Janssen
Dragana
D
Spasic
Jeroen
J
Heyligen
Kim
K
Vriens
Karin
K
Thevissen
Bruno P A
BP
Cammue
Valery
V
Petrenko
Chris
C
Ulens
Hans
H
Deckmyn
Jeroen
J
Lammertyn
D001435:Bacteriophages / Q000235:genetics*; D002352:Carrier Proteins / Q000032:analysis* / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D049452:Green Fluorescent Proteins / Q000737:chemistry; D036341:Intercellular Signaling Peptides and Proteins; D010455:Peptides / Q000032:analysis* / Q000235:genetics; D020349:Surface Plasmon Resonance / Q000379:methods*
D002352:Carrier Proteins; D036341:Intercellular Signaling Peptides and Proteins; C541129:P8 peptide; D010455:Peptides; C485184:enhanced green fluorescent protein; C485059:peptide P3; D049452:Green Fluorescent Proteins