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Mechanism of the biosynthesis of carbon monoxide necessary for the expression of the metalloenzyme activity (Press Release)

Release Date
07 Nov, 2019
  • BL44XU (Macromolecular Assemblies)

Thursday 7 November 2019
Institute for Molecular Science

The research group including researchers of National Institutes of Natural Sciences (ExCELLS/IMS), and Osaka University revealed the detail mechanism of the biosynthesis of carbon monoxide essential for the maturation of the unit responsible for hydrogenase activity.

The findings have been published in Communications Biology (Nature Research).

Journal Article
Title:Structural characterization of HypX responsible for CO biosynthesis in the maturation of NiFe-hydrogenase
Authors:Norifumi Muraki, Kentaro Ishii, Susumu Uchiyama, Satoru G. Itoh, Hisashi Okumura, Shigetoshi Aono
Journal:Communications Biology(October 18, 2019 Online)

Hydrogenase, which is a metalloenzyme responsible for the oxidation of hydrogen gas and the reduction of proton, plays a key role of bacterial hydrogen metabolism. Based on the differences of metal content of the active site, they are classified into three groups; FeFe-, NiFe-, and Fe-hydrogenases containing different metal complexes as active centers in these enzymes (Figure 1). Although the structures of the active centers in these three enzymes are different, it is essential for hydrogenase activity that carbon monoxide (CO) is coordinated to the iron ion in the active center. It’s known that CO was synthesized by an enzymatic reaction, but the detail of CO biosynthesis was unknown.

Figure 1..

Figure 1. Structure of the active sites in [NiFe]-hydrogenase, [FeFe]-hydrogenase, and [Fe]-hydrogenase

In this research, the research group determined the crystal structure of the enzyme (HypX) responsible for the biosynthesis of CO in which NiFe-hydrogenase utilize and revealed that HypX produce CO by unprecedented reaction (Figure 2). HypX consists of two domains; the N-terminal and C-terminal domain. A continuous cavity connecting the N- and C-terminal domains is present in the interior of HypX (Figure 2). In the crystal structure, coenzyme A (CoA) is bound in the C-terminal region of the cavity.

Figure 2

Figure 2. Crystal structure of HypX The N- and C-terminal domains are shown in blue and green respectively. The cavity in side of HypX is shown in grey mesh.

Two different reactions occur in the N- and C-terminal domain, respectively. In the N-terminal domain, formyl-group transfer reaction from formyltetrahydrofolate, which is bound in the N-terminal region of the cavity as a substrate to CoA takes place (reaction step 1 in Fig. 3). At this time, CoA in the cavity adopts the linear extended conformation, and the SH-group of CoA is located on the side of the formyl group in the formyltetrahydrofolate bonded to the N-terminal domain. Then, formyl-CoA is produced as a reaction intermediate by the formyl-group transfer reaction from formyltetrahydrofolate to CoA. Next, formyl-CoA undergoes a large conformational change in the cavity so that the formyl group in the terminal portion of formyl-CoA is located at the active site in the C-terminal domain of HypX (reaction step 2 in Fig. 3). In the C-terminal domain, CO and CoA are formed by decarbonylation of formyl-CoA (reaction step 3 in Fig. 3).

Figure 3

Figure 3. Reaction scheme of CO biosynthesis by HypX The N- and C-terminal domains are shown in blue and green respectively. Formyl-group (CHO) is shown in red.

In this study, we revealed the unprecedented CO synthesis reaction. CoA is well known as a coenzyme, which has the important role in fatty acid metabolism and cellular energetic metabolism through the citric acid cycle. However, it has never been reported that CoA (formyl-CoA) is involved in CO biosynthesis reactions. This study clarified the novel physiological function of a well-known coenzyme CoA.

Future prospects
The biosynthetic mechanism of metalloenzyme has still many unclear points. It remains to be elucidated especially how the complex metal-containing active center of metalloenzyme was assembled. In this work, we determined the first crystal structure of the enzyme, which catalyzes the biosynthetic reaction of carbon monoxide essential for the construction of the active site of [NiFe] hydrogenase. In future, we will continue the research for elucidating the detail mechanism of whole hydrogenase maturation pathway based on this result.

A Grant-in-Aid for Scientific Research (B) 17H03093 Joint Research by Exploratory Research Center on Life and Living Systems (ExCELLS).

Shigetoshi Aono
Exploratory Research Center on Life and Living Systems (ExCELLS)/
Institute for Molecular Science, Natural Institutes of Natural Sciences
 TEL: +81-564-59-5575 / FAX: +81-564-59-5576
 E-mail: aonoatims.ac.jp

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