Mechanistic Insight into the Enzymatic Reduction of Truncated Hemoglobin N of Mycobacterium tuberculosis

Singh, Sandeep; Thakur, Naveen; Oliveira, Ana; Petruk, Ariel Alcides; Hade, Mangesh Dattu; Sethi, Deepti; Bidon‐Chanal, Axel; Martí, Marcelo A.; Datta, Himani; Parkesh, Raman; Estrin, Darı́o A.; Luque, F. Javier; Dikshit, Kanak L.

doi:10.1074/jbc.m114.578187

Cited by 15 publications

(14 citation statements)

References 46 publications

(85 reference statements)

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“…Herein we have shown that THB1 is very efficiently reduced by NR, which contains FAD as a cofactor (Figure 4d). It is noteworthy that GlbN from Mycobacterium tuberculosis can be reduced in vitro at a concentration and velocity similar to those of CrTHB1 by the NADH ferredoxin/flavodoxin reductase system from E. coli (Singh et al, 2014). NR takes electrons from NAD(P)H to reduce nitrate or alternatively to transfer these electrons to other acceptors from FAD (diaphorase activity).…”

Section: Discussionmentioning

confidence: 99%

THB1, a truncated hemoglobin, modulates nitric oxide levels and nitrate reductase activity

et al. 2015

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These authors contributed equally to this work. SUMMARYHemoglobins are ubiquitous proteins that sense, store and transport oxygen, but the physiological processes in which they are implicated is currently expanding. Recent examples of previously unknown hemoglobin functions, which include scavenging of the signaling molecule nitric oxide (NO), illustrate how the implication of hemoglobins in different cell signaling processes is only starting to be unraveled. The extent and diversity of the hemoglobin protein family suggest that hemoglobins have diverged and have potentially evolved specialized functions in certain organisms. A unique model organism to study this functional diversity at the cellular level is the green alga Chlamydomonas reinhardtii because, among other reasons, it contains an unusually high number of a particular type of hemoglobins known as truncated hemoglobins (THB1-THB12). Here, we reveal a cell signaling function for a truncated hemoglobin of Chlamydomonas that affects the nitrogen assimilation pathway by simultaneously modulating NO levels and nitrate reductase (NR) activity. First, we found that THB1 and THB2 expression is modulated by the nitrogen source and depends on NIT2, a transcription factor required for nitrate assimilation genes expression. Furthermore, THB1 is highly expressed in the presence of NO and is able to convert NO into nitrate in vitro. Finally, THB1 is maintained on its active and reduced form by NR, and in vivo lower expression of THB1 results in increased NR activity. Thus, THB1 plays a dual role in NO detoxification and in the modulation of NR activity. This mechanism can partly explain how NO inhibits NR post-translationally.

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Section: Discussionmentioning

confidence: 99%

THB1, a truncated hemoglobin, modulates nitric oxide levels and nitrate reductase activity

et al. 2015

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“…The microbe-derived HbN decomposes NO produced by macrophages and neutrophils so the bacterium may survive within the cytosol. HbN acts as a NO dioxygenase despite the absence of a true reductase domain [115]. HbN expression intensifies once the bacterium enters the WBC.…”

Section: Use Of Hemoglobin By Microbesmentioning

confidence: 99%

Immunological properties of oxygen-transport proteins: hemoglobin, hemocyanin and hemerythrin

Coates

Decker

2016

Cell. Mol. Life Sci.

119

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It is now well documented that peptides with enhanced or alternative functionality (termed cryptides) can be liberated from larger, and sometimes inactive, proteins. A primary example of this phenomenon is the oxygen-transport protein hemoglobin. Aside from respiration, hemoglobin and hemoglobin-derived peptides have been associated with immune modulation, hematopoiesis, signal transduction and microbicidal activities in metazoans. Likewise, the functional equivalents to hemoglobin in invertebrates, namely hemocyanin and hemerythrin, act as potent immune effectors under certain physiological conditions. The purpose of this review is to evaluate the true extent of oxygen-transport protein dynamics in innate immunity, and to impress upon the reader the multi-functionality of these ancient proteins on the basis of their structures. In this context, erythrocyte–pathogen antibiosis and the immune competences of various erythroid cells are compared across diverse taxa.Electronic supplementary materialThe online version of this article (doi:10.1007/s00018-016-2326-7) contains supplementary material, which is available to authorized users.

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“…O 2 and NO tunnels and a nitrate egress pathway have been suggested to play roles in the NOD function of the truncated HbN from Mycobacterium tuberculosis ( 22 , 23 ). A long hydrophobic tunnel that runs parallel to the H helix and perpendicular to the heme plane is thought to allow NO ( 22 , 24 , 25 ), or O 2 ( 16 , 24 , 26 ), to access the distal heme reaction chamber, while a short tunnel formed between the G and H helices at the AlaG5 and LeuH8 residues allows O 2 ( 22 ), or NO ( 24 , 26 , 27 ), to access the distal heme pocket. Moreover, movement of PheE15 caused by O 2 binding to the ferrous heme and TyrB10-GlnE11 hydrogen-bonding interactions ( 28 ) may function as a gate, in cooperation with LeuG8, that controls NO access to the heme ( 22 , 25 , 29 , 30 , 31 ).…”

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confidence: 99%

Allostery in the nitric oxide dioxygenase mechanism of flavohemoglobin

Gardner

2021

Journal of Biological Chemistry

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The substrates O 2 and NO cooperatively activate the NO dioxygenase function of Escherichia coli flavohemoglobin. Steady-state and transient kinetic measurements support a structure-based mechanistic model in which O 2 and NO movements and conserved amino acids at the E11, G8, E2, E7, B10, and F7 positions within the globin domain control activation. In the cooperative and allosteric mechanism, O 2 migrates to the catalytic heme site via a long hydrophobic tunnel and displaces LeuE11 away from the ferric iron, which forces open a short tunnel to the catalytic site gated by the ValG8/IleE15 pair and LeuE11. NO permeates this tunnel and leverages upon the gating side chains trigger the CD loop to furl, which moves the E and F-helices and switches an electron transfer gate formed by LysF7, GlnE7, and water. This allows FADH 2 to reduce the ferric iron, which forms the stable ferric–superoxide–TyrB10/GlnE7 complex. This complex reacts with internalized NO with a bimolecular rate constant of 10 10 M −1 s −1 forming nitrate, which migrates to the CD loop and unfurls the spring-like structure. To restart the cycle, LeuE11 toggles back to the ferric iron. Actuating electron transfer with O 2 and NO movements averts irreversible NO poisoning and reductive inactivation of the enzyme. Together, structure snapshots and kinetic constants provide glimpses of intermediate conformational states, time scales for motion, and associated energies.

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Mechanistic Insight into the Enzymatic Reduction of Truncated Hemoglobin N of Mycobacterium tuberculosis

Cited by 15 publications

References 46 publications

THB1, a truncated hemoglobin, modulates nitric oxide levels and nitrate reductase activity

THB1, a truncated hemoglobin, modulates nitric oxide levels and nitrate reductase activity

Immunological properties of oxygen-transport proteins: hemoglobin, hemocyanin and hemerythrin

Allostery in the nitric oxide dioxygenase mechanism of flavohemoglobin

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