Stepwise Hydride Transfer in a Biological System: Insights into the Reaction Mechanism of the Light‐Dependent Protochlorophyllide Oxidoreductase

Archipowa, Nataliya; Kutta, Roger Jan; Heyes, Derren J.; Scrutton, Nigel S.

doi:10.1002/ange.201712729

Cited by 21 publications

(44 citation statements)

References 33 publications

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“…As a specialized SDR, LPOR catalyzes the light-dependent reduction of Pchlide. Its catalytic mechanism has been investigated on the micro-to picosecond timescales (1,16,24,26,33,35,47). The process can be briefly described as: The photoactivated Pchlide receives a hydride from the electron donor NADPH at C17 of the C17 = C18 double bond, then a proton is transferred through a conserved tyrosine (Tyr193 in SyLPOR and TeLPOR) to C18, resulting in two chiral centers at C17 and C18.…”

Section: Discussionmentioning

confidence: 99%

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Crystal structures of cyanobacterial light-dependent protochlorophyllide oxidoreductase

Dong

Zhang

Wang

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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The reduction of protochlorophyllide (Pchlide) to chlorophyllide (Chlide) is the penultimate step of chlorophyll biosynthesis. In oxygenic photosynthetic bacteria, algae, and plants, this reaction can be catalyzed by the light-dependent Pchlide oxidoreductase (LPOR), a member of the short-chain dehydrogenase superfamily sharing a conserved Rossmann fold for NAD(P)H binding and the catalytic activity. Whereas modeling and simulation approaches have been used to study the catalytic mechanism of this light-driven reaction, key details of the LPOR structure remain unclear. We determined the crystal structures of LPOR from two cyanobacteria, Synechocystis sp. PCC 6803 and Thermosynechococcus elongatus. Structural analysis defines the LPOR core fold, outlines the LPOR–NADPH interaction network, identifies the residues forming the substrate cavity and the proton-relay path, and reveals the role of the LPOR-specific loop. These findings provide a basis for understanding the structure-function relationships of the light-driven Pchlide reduction.

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Section: Discussionmentioning

confidence: 99%

“…Extensive biochemical and biophysical studies have been performed on LPOR from two cyanobacteria, S. sp. PCC 6803 (18)(19)(20)(21)(22)(23)(24)(25)(26) and T. elongatus (27)(28)(29)(30)(31)(32)(33)(34)(35). The catalytic process has been demonstrated to be initiated by the absorption of light.…”

mentioning

confidence: 99%

Crystal structures of cyanobacterial light-dependent protochlorophyllide oxidoreductase

Dong

Zhang

Wang

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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“…Nature has discovered two completely different ways to achieve this reduction: in one, a lightdependent reaction is catalyzed by NADPH:protochlorophyllide oxidoreductase (LPOR; EC 1.3.1.33) (207,208); in the other, reduction of the C17-C18 double bond is catalyzed by a dark-operative protochlorophyllide reductase (DPOR), consisting of ChlL, ChlN and ChlB subunits that display similarity to the components of nitrogenase (209). The ability to trigger the catalytic cycle with short pulses of light has led to a number of kinetic studies (210)(211)(212), and recently the structure of LPOR has been reported (208). The phylogenetic distribution of LPOR and DPOR is interesting; anoxygenic photosynthetic bacteria contain only DPOR; cyanobacteria, green algae, mosses and most gymnosperms possess both LPOR and DPOR; and angiosperms (flowering plants) contain only LPOR.…”

Section: The Transformation Of Protoix Into Chls -Chl Amentioning

confidence: 99%

Biosynthesis of the modified tetrapyrroles—the pigments of life

Bryant

Hunter

Warren

2020

Journal of Biological Chemistry

188

164

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Modified tetrapyrroles are large macrocyclic compounds, consisting of diverse conjugation and metal chelation systems and imparting an array of colors to the biological structures that contain them. Tetrapyrroles represent some of the most complex small molecules synthesized by cells and are involved in many essential processes that are fundamental to life on Earth, including photosynthesis, respiration, and catalysis. These molecules are all derived from a common template through a series of enzyme-mediated transformations that alter the oxidation state of the macrocycle and also modify its size, its side-chain composition, and the nature of the centrally chelated metal ion. The different modified tetrapyrroles include chlorophylls, hemes, siroheme, corrins (including vitamin B12), coenzyme F430, heme d1, and bilins. After nearly a century of study, almost all of the more than 90 different enzymes that synthesize this family of compounds are now known, and expression of reconstructed operons in heterologous hosts has confirmed that most pathways are complete. Aside from the highly diverse nature of the chemical reactions catalyzed, an interesting aspect of comparative biochemistry is to see how different enzymes and even entire pathways have evolved to perform alternative chemical reactions to produce the same end products in the presence and absence of oxygen. Although there is still much to learn, our current understanding of tetrapyrrole biogenesis represents a remarkable biochemical milestone that is summarized in this review.

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“…Pchlide excited-state electron transfer from NADPH followed by a proton-coupled electron transfer, 18 which represents the first example of the stepwise transfer of a hydride equivalent reported in biology. These time-resolved studies have provided insight into the chemistry of catalysis across a wide range of timescales (from the femtosecond to the second), but the required structural basis of POR photocatalysis has remained unknown.…”

mentioning

confidence: 99%

“…10,24,26 These excited-state interactions between Pchlide and POR stabilize a highly polarized C-17-C-18 double bond and enables stepwise hydride transfer from NADPH to C-17. 18 The polarized nature of the C-17-C-18 bond may also be stabilized by the close proximity of the hydroxyl group of Tyr193, which is known to be required for Pchlide photochemistry. 10 By contrast, Tyr223 does not form any direct interaction with Pchlide (Extended Data Fig.…”

mentioning

confidence: 99%

Structural basis for enzymatic photocatalysis in chlorophyll biosynthesis

Zhang

Heyes

Feng

et al. 2019

Nature

Self Cite

109

190

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The enzyme protochlorophyllide oxidoreductase (POR) catalyses a lightdependent step in chlorophyll biosynthesis that is essential to photosynthesis and ultimately all life on Earth. 1-3 POR, which is one of three known light-dependent enzymes, 4,5 catalyzes reduction of the photosensitizer and substrate protochlorophyllide to form the pigment chlorophyllide. Despite its biological importance, a structural basis for POR photocatalysis has remained elusive. Here, we report crystal structures of cyanobacterial PORs from Thermosynechococcus elongatus and Synechocystis sp. in their free forms, and in complex with nicotinamide coenzyme. Our structural models and simulations of the ternary protochlorophyllide-NADPH-POR complex have identified multiple interactions in the POR active site that are important for protochlorophyllide binding, photosensitization and photochemical conversion to chlorophyllide. We demonstrate the importance of active-site architecture and protochlorophyllide structure in experiments using POR variants and protochlorophyllide analogues. These studies reveal how the POR active site facilitates light-driven reduction of protochlorophyllide by localized hydride transfer from NADPH and long-range proton transfer along structurally defined proton-transfer pathways. As the light-driven step in the chlorophyll biosynthetic pathway (Fig. 1), the POR reaction acts as the trigger for the germination of seedlings =in plants and provokes a marked change in the morphological development of the plant. 2,3 Given this crucial biological role, POR has been the focus of numerous mechanistic and biophysical investigations. A combination of time-resolved (at the femtosecond-to-second scale) and cryogenic spectroscopy methods have provided some understanding of the mechanism of POR photocatalysis in a range of photosynthetic organisms, including cyanobacteria and plants. Picosecond excited-state dynamics in the protochlorophyllide (Pchlide) molecule are thought to result in excited state interactions between the substrate and active-site residues that are necessary to trigger the subsequent reaction chemistry. 6-12 This involves sequential transfer of a hydride equivalent from NADPH and a proton transfer from either an active site residue or solvent. Proton transfer is reliant on solvent dynamics and an implied network of extended protein motions that occur on the microsecond timescale. 13-17 Hydride transfer from NADPH is not concerted, but occurs in a stepwise manner that involves

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Stepwise Hydride Transfer in a Biological System: Insights into the Reaction Mechanism of the Light‐Dependent Protochlorophyllide Oxidoreductase

Cited by 21 publications

References 33 publications

Crystal structures of cyanobacterial light-dependent protochlorophyllide oxidoreductase

Crystal structures of cyanobacterial light-dependent protochlorophyllide oxidoreductase

Biosynthesis of the modified tetrapyrroles—the pigments of life

Structural basis for enzymatic photocatalysis in chlorophyll biosynthesis

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