(27) Assay Card

General Information
Summary RTD-1-3 shown to bind with equal affinity to E.coli.
Condition Binding of RTD-1-3 to E.¬coli ML35 was evaluated by incubating 2¬ó6 log-phase bacteria with increasing peptide concentrations (0.5-8 ¬µg/ml final) in 1¬ml of 10¬mM PIPES, pH 7.4,¬containing 5¬mM glucose for 2¬h at 37¬¬C. The incubation mixtures were centrifuged at 20,000-25,000¬ófor 10¬min at 22¬¬C, and supernatant samples were analyzed for RTD content by RP-HPLC. Binding of each RTD to E.¬coli was determined by subtraction of the quantity of supernatant peptide from total added to each tube, and comparing this to control incubations lacking bacteria.
Result The binding of the three theta-defensins to E.¬coli cells was essentially identical. For all three theta-defensins, maximal peptide binding by 2¬ó6 bacteria was ~2 ug.
AssayType Membrane Binding Assay

Tran D, Tran PA, Tang YQ, Yuan J, Cole T, Selsted ME (2002) Homodimeric theta-defensins from rhesus macaque leukocytes: isolation, synthesis, antimicrobial activities, and bacterial binding properties of the cyclic peptides. J Biol Chem 277:3079-84

Proteins Assayed RTD-1