Structural Basis for the Regulation of Biofilm Formation and Iron Uptake in <i>A. baumannii</i> by the Blue-Light-Using Photoreceptor, BlsA

Chitrakar, Iva; Iuliano, James N.; He, Yong Le; Woroniecka, Helena A.; Collado, Jinnette Tolentino; Wint, Jinelle M.; Walker, Stephen G.; Tonge, Peter J.; French, Jarrod B.

doi:10.1021/acsinfecdis.0c00156

Cited by 17 publications

(24 citation statements)

References 61 publications

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“…This fact goes in good agreement with structural predictions from models of these two proteins generated using the crystallographic structure of dark- and light-adapted states of BlsA (PDBs 6W6Z and 6W72, respectively) as templates (Fig 5). Fig 5A and 5B show the location of water molecules (blue dots) for the dark-adapted form of BlsA (d-BlsA) and the light-adapted (l-BlsA) respectively, determined by Chitrakar et al [32]. The flavin pocket access site has more affinity and/or accessibility to water molecules in the light-adapted form, supporting our previous characterization of chromophore behavior inside BlsA cavity by the flavin fluorescence emission (Fig 5A and B)[10].…”

Section: Resultsmentioning

confidence: 99%

Characterization of BLUF-photoreceptors present in Acinetobacter nosocomialis

Abatedaga

Mora

Tuttobene

et al. 2021

Preprint

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Acinetobacter nosocomialis is a Gram-negative opportunistic pathogen, whose ability to cause disease in humans is well recognized. Blue light has been shown to modulate important physiological traits related to persistence and virulence in this microorganism. In this work, we characterized the three Blue Light sensing Using FAD (BLUF) domain-containing proteins encoded in the A. nosocomialis genome, which account for the only "traditional" light sensors present in this microorganism. By focusing on a light-modulated bacterial process such as motility, the temperature dependence of light regulation was studied, as well as the expression pattern and spectroscopic characteristics of the different A. nosocomialis BLUFs. Our results show that the three BLUF-containing proteins encode active photoreceptors, despite only two of them are stable in the light-regulatory temperature range when expressed recombinantly. In vivo, only the A. baumannii's ortholog AnBLUF65 is expressed, which is active in a temperature range from 15 °C to 37 °C. In turn, AnBLUF46 is an active photoreceptor between 15 °C to 32 °C in vitro, but is not expressed in A. nosocomialis in the conditions tested. Intra-protein interactions were analyzed using 3D models built based on A. baumanni's photoreceptor, to support spectroscopic data and profile intra-protein residue interactions. A general scheme is presented on how hydrophobic/aromatic interactions may contribute to the stability of dark/light- adapted states, indicating the importance of these interactions in BLUF photoreceptors.

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Section: Resultsmentioning

confidence: 99%

Characterization of BLUF-photoreceptors present in Acinetobacter nosocomialis

Abatedaga

Mora

Tuttobene

et al. 2021

Preprint

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“…Recent work involving the crystallization of BlsA, protein docking prediction experiments, and binding-affinity analyses has identified particular BlsA residues that are necessary for its interaction with Fur. Indeed, residues 101-110 are required for this interaction ( Chitrakar et al., 2020 ). Although the significance of BlsA’s last five amino acid residues was not addressed in the study by Chitrakar et al.…”

Section: Discussionmentioning

confidence: 99%

“…Indeed, residues 101-110 are required for this interaction ( Chitrakar et al., 2020 ). Although the significance of BlsA’s last five amino acid residues was not addressed in the study by Chitrakar et al. (2020) , primarily because of their unsolvable nature during crystallography, further protein-protein interaction analyses would elucidate whether BlsA’s last five residues are necessary for stabilization of its interaction with the Fur transcriptional regulator.…”

Section: Discussionmentioning

confidence: 99%

The Influence of Blue Light and the BlsA Photoreceptor on the Oxidative Stress Resistance Mechanisms of Acinetobacter baumannii

Squire

Townsend

Actis

2022

Front. Cell. Infect. Microbiol.

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Acinetobacter baumannii is a catalase-positive Gram-negative bacterial pathogen that causes severe infections among compromised patients. Among its noteworthy regulatory mechanisms, this microorganism regulates its lifestyle through the blue light using flavin (BLUF) protein BlsA. This protein regulates a diverse set of cellular processes that include, but are not limited to, motility, biofilm formation, phenylacetic acid metabolism, iron uptake, and catalase activity. We set out to determine how A. baumannii regulates catalase activity and other related oxidative stress phenotypes in response to light. Notably, because A. baumannii ATCC 17978 encodes four catalase homologs – which we refer to as KatA, KatE, KatE2, and KatG – we also aimed to show which of these enzymes exhibit light- and BlsA-dependent activity. Our work not only provides insight into the general function of all four catalase homologs and the impact of light on these functions, but also directly identifies KatE as a BlsA-regulated enzyme. We further demonstrate that the regulation of KatE by BlsA is dependent on a lysine residue that we previously demonstrated to be necessary for the regulation of surface motility. Furthermore, we show that BlsA’s five most-C-terminal residues – previously considered dispensable for BlsA’s overall function – are necessary for the light-independent and light-dependent regulation of catalase and superoxide dismutase activities, respectively. We hypothesize that these identified critical residues are necessary for BlsA’s interaction with protein partners including the transcriptional regulators Fur and BfmR. Together these data expand the understanding regarding how A. baumannii uses light as a signal to control oxidative stress resistance mechanisms that are critical for its pathophysiology.

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“…NMR measurements of BlrP1 indicate that the blue light induces conformational changes in α3-and α4-helices extending from the Cterminus of the BLUF domain. 39 In the case of BlrP1, the C-terminal cap helix is docked on the strands of the BLUF domain, like in other BLUF proteins, such as PixDs, 4,6 AppA, 34 OaPAC, 9 bPAC, 40 BlrB, 5 and BlsA, 41 but the orientation of the helix relative to the -strands is different in BlrP1 from the others (parallel for BlrP1 and perpendicular for the others). 42 As a result, the distance between the position of Trp, which is replaced by Thr in BlrP1, and the cap helix is longer in BlrP1 than the other proteins.…”

Section: Origin Of τ 2 -Phasementioning

confidence: 99%

Slow conformational changes of blue light sensor BLUF proteins in milliseconds

Tokonami

Onose

Nakasone

et al. 2021

Preprint

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BLUF (blue light sensor using flavin) proteins consist of flavin-binding BLUF domains and functional domains. Upon blue light excitation, the hydrogen bond network around the flavin chromophore changes, and the absorption spectrum in the visible region exhibits red-shift. Ultimately, the light information received in the BLUF domain is transmitted to the functional region. It has been believed that this red-shift is complete within nanoseconds. Contrary to this commonly accepted scheme, in this study, slow reaction kinetics were discovered in milliseconds (τ1- and τ2-phase) for all the BLUF proteins examined (AppA, OaPAC, BlrP1, YcgF, PapB, SyPixD, and TePixD). Despite extensive reports on BLUF, this is the first clear observation of the BLUF protein absorption change with the duration in the millisecond time region. From the measurements of some domain-deleted mutants of OaPAC and two chimeric mutants of PixD proteins, it was found that the slower dynamics (τ2-phase) are strongly affected by the size and nature of the C-terminal region adjacent to the BLUF domain. Hence, this millisecond reaction is a significant indicator of conformational changes in the C-terminal region, which is essential for the biological functions. On the other hand, the τ1-phase commonly exists in all BLUF proteins, including any mutants. The origin of the slow dynamics was studied using site-specific mutants. These results clearly show the importance of Trp in the BLUF domain. Based on this, a reaction scheme for the BLUF reaction is proposed.

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Structural Basis for the Regulation of Biofilm Formation and Iron Uptake in A. baumannii by the Blue-Light-Using Photoreceptor, BlsA

Cited by 17 publications

References 61 publications

Characterization of BLUF-photoreceptors present in Acinetobacter nosocomialis

Characterization of BLUF-photoreceptors present in Acinetobacter nosocomialis

The Influence of Blue Light and the BlsA Photoreceptor on the Oxidative Stress Resistance Mechanisms of Acinetobacter baumannii

Slow conformational changes of blue light sensor BLUF proteins in milliseconds

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