کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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1933980 | 1050631 | 2009 | 5 صفحه PDF | دانلود رایگان |
The binding affinity of the N-terminal peptidoglycan binding domain of endolysin KZ144 (PBDKZ), originating from Pseudomonas aeruginosa bacteriophage ϕKZ, has been examined using a fusion protein of PBDKZ and green fluorescent protein (PBDKZ-GFP). A fluorescence recovery after photobleaching analysis of bound PBDKZ-GFP molecules showed less than 10% fluorescence recovery in the bleached area within 15 min. Surface plasmon resonance analysis confirmed this apparent high binding affinity revealing an equilibrium affinity constant of 2.95 × 107 M−1 for the PBDKZ-peptidoglycan interaction. This unique domain, which binds to the peptidoglycan of all tested Gram-negative species, was harnessed to improve the specific activity of the peptidoglycan hydrolase domain KMV36C. The chimeric peptidoglycan hydrolase (PBDKZ-KMV36C) exhibits a threefold higher specific activity than the native catalytic domain (KMV36C). These results demonstrate that the modular assembly of functional domains is a rational approach to improve the specific activity of endolysins from phages infecting Gram-negatives.
Journal: Biochemical and Biophysical Research Communications - Volume 383, Issue 2, 29 May 2009, Pages 187–191