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Photo-dynamics of BLUF domain containing adenylyl cyclase NgPAC3 from the amoeboflagellate Naegleria gruberi NEG-M strain

Paper ID Volume ID Publish Year Pages File Format Full-Text
27016 43994 2014 11 PDF Available
Title
Photo-dynamics of BLUF domain containing adenylyl cyclase NgPAC3 from the amoeboflagellate Naegleria gruberi NEG-M strain
Abstract

•Photo-activated adenylyl cyclase NgPAC3 from Naegleria gruberi NEG-M was expressed.•Photo-cycle dynamics of BLUF domain in NgPAC3 protein was studied.•Flavin cofactor is partly fully oxidized and partly fully reduced.•Adenylyl cyclase activity was studied under dark and light adapted conditions.•Thermal stability was studied and apparent melting temperature was determined.

The absorption and emission spectroscopic behavior of the photo-activated adenylyl cyclase NgPAC3 from the amoeboflagellate Naegleria gruberi NEG-M strain was studied. The flavin cofactor was found to be partly fully oxidized and partly fully reduced. The typical BLUF domain (BLUF = Blue Light sensor Using Flavin) oxidized flavin absorption photo-cycle dynamics with about 14 nm flavin absorption red-shift in the signaling state was observed. The quantum efficiency of signaling state formation was determined to be ϕs = 0.66 ± 0.03. A bi-exponential signaling state recovery to the dark-adapted receptor state was observed with the time constants τrec,f = 275 s and τrec,sl = 45 min. The thermal irreversible protein unfolding was studied and an apparent protein melting temperature of ϑm ≈ 50 °C was found. The photodynamic behavior of NgPAC3 is compared with the behavior of the previously investigated photo-activated cyclases NgPAC1 (nPAC) and NgPAC2 from the same N. gruberi NEG-M strain. Purified recombinant NgPAC3 showed light-gated adenylate cyclase activity upon illumination with blue light. Its cyclase activity is compared with those of NgPAC1 and NgPAC2.

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Keywords
Photo-activated cyclase; BLUF domain; Amoeboflagellate Naegleria gruberi; Photo-cycle dynamics; Apparent protein melting temperature; Adenylyl cyclase activity
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Photo-dynamics of BLUF domain containing adenylyl cyclase NgPAC3 from the amoeboflagellate Naegleria gruberi NEG-M strain
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Photochemistry and Photobiology A: Chemistry - Volume 287, 1 August 2014, Pages 19–29
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us