Methylmalonate Semialdehyde Dehydrogenase (MMSDH) in Oceanimonas doudoroffii

Megan Gothard '19 and Andrea Ludwig '19


I. Introduction

Oceanimonas doudoroffii is a gram-negative bacterium found in seawater. 1 Its primary carbon and sulfur source is dimethylsulfoniopropionate (DMSP), a compound produced by halophytes and algaes to maintain osmotic pressures during times of stress. 2 The DMSP demethylation pathway and the DMSP cleavage pathway compete to break down DMSP within Oceanimonas doudoroffii. 1

Within the DMSP cleavage pathway is the enzyme methylmalonate-semialdehyde dehydrogenase (MMSDH). 1 MMSDH catalyzes the decarboxylation of the intermediate methylmalonate semialdehyde (MMSA) to propionyl-CoA downstream in the DMSP pathway. 1 To complete this reaction, MMSDH requires CoA and NAD + cofactors. NAD + is hydrogenated by MMSDH to convert MMSA into propionyl-CoA (Figure 1). 1

To better explain this process, we will examine the structure of MMSDH and delve into catalytic domains that make up the monomeric structure of MMSDH: the NAD-binding domain and the substrate-binding domain. Before we focus on the binding domains, however, we will first consider the overall structure of MMSDH.

II. General Protein Structure

The structure of MMSDH is formed by six in solution. The six dimers show little structural variation, so the catalytic activity can be more easily understood by considering one representative dimer, . Breaking down the structure further, Dimer A of MMSDH is comprised of a total of two monomers, which are together capable of binding an NAD molecule. Each is composed of 14 alpha-helices and 19 beta-strands, and contain the three of MMSDH: the NAD-binding domain, the substrate binding domain, and the oligomerization domain. The NAD-binding domain and substrate binding domains are composed of six central beta sheets surrounded by alpha helices. The oligomerization domain is composed of three antiparallel beta sheets. The third domain, the oligomerization domain, serves only to connect the monomers of MMSDH, serves no catalytic function, and will not be further highlighted.  

III. NAD-Binding Domain

The is composed of 6 alpha helices (a2-a7, a14) and 7 beta sheets (B1-B4, B7-B11). In the presence of an , monomers in the MMSDH molecule bind NAD + and hydrogenate it, to form NADH. This reduced form of NAD + is then able to accept an electron from MMSA, which allows for its conversion to propionyl CoA. The NAD + binding region of MMSDH binds NAD + via hydrophobic interactions and hydrogen bonding.

The hydrophobic adenine ring of NAD + interacts with the hydrophobic pocket of MMSDH formed by Val208 and Ile228. Also, the nicotinamide ring of NAD + forms hydrophobic interactions with Val223, Gly222, Pro147, and Val154, of the NAD binding domain.

The adenine ribose ring of NAD + forms hydrogen bonds with the side chain of Lys172 and the phosphate group of NAD + forms a hydrogen bond with Ser225. Additionally, the nicotinamide ribose oxygen of NAD + hydrogen bonds with both Gly247 and Glu382.

IV. Substrate Binding Domain

The is made up of numerous amino acid residues that make the environment favorable for MMSA to bind. The substrate binding domain is located very close to the NAD-binding sites, allowing the domains to conformationally promote the hydrogenation of NAD + and the protonation of MMSA. However, the entrances to these sites are in opposite locations and the two binding sites are in opposite locations. The of MMSDH is surrounded by residues Arg103, Arg279, Leu440, Ser275, and Val442. These amino acids line the bottom of the pocket with functional groups, which interact with the polar regions of MMSA. Arg103 and Arg279 likely anchor the negative C-1 carboxyl group of MMSA. The positive environment of the binding site makes the interactions between the substrate and MMSDH favorable. The catalytic residue serves as a proton donor for the catalytic conversion of NAD + to NADH, providing the catalytic capabilities of MMSDH to produce propionyl-CoA.  

V. Implications of MMSDH in Humans

Interestingly, the MMSDH enzyme is conserved between Oceanimonas doudoroffii and humans. 3 Within humans, MMSDH is responsible for branched chain amino acid (BCAA) degradation. 3 BCAAs are essential amino acids in humans, and must be consumed for normal development to occur. 4 As can be expected, mutations in the enzymes of the valine degradation pathway have rare but serious health effects. 3 Patients with homologous point mutations in the MMSDH gene (ALDH6A1) have severe developmental delays and high levels of 3-hydroxyisobutyric acid (shown in red below). 3 This valine degradation intermediate is caused by the lack of MMSDH functionality. 3The buildup of acid intermediates causes severe ketoacidosis (low blood pH) and often results in early childhood fatality. 3

Patients with less serious mutations in the BCAA degradation pathway often have Maple Syrup Urine Disease (MSUD).5 The characteristic symptoms of MSUD are bodily fluids that smell sweet like maple syrup. 5 This characteristic scent is caused by BCAA degradation intermediate accumulation. 5 With early detection of this disease, patients' diets can reduce leucine, isoleucine, and valine from their diets before toxic intermediates accumulate and symptoms worsen and become deadly. 5 In 2007, infant blood testing for MSUD has been mandated in the United States so as to prevent the accumulation of BCAAs that causes such serious symptoms in these affected individuals. 5

VI. References

1. Do, Hackwon, Chang Woo Lee, Sung Gu Lee, Hara Kang, Chul Min Park, Hak Jun Kin, Hyun Park, HaJeung Park, Jun Hyuck Lee. Crystal structure and modeling of the tetrahedral intermediate state of methylmalonate-semialdehyde dehydrogenase (MMSDH) from Oceanimonas doudoroffii. Journal of Microbiology. 54, 114–121 (2016).

2. Yoch, D. C. Dimethylsulfoniopropionate: its sources, role in the marine food web, and biological degradation to dimethylsulfide. Appl. Environ. Microbiol. 68, 5804–15 (2002).

3.Sass, J. O. et al. 3-Hydroxyisobutyrate aciduria and mutations in the ALDH6A1 gene coding for methylmalonate semialdehyde dehydrogenase. J. Inherit. Metab. Dis. 35, 437–442 (2012).

4.Hutson, S. M., Sweatt, A. J. & LaNoue, K. F. Branched-Chain Amino Acid Metabolism: Implications for Establishing Safe Intakes. J. Nutr. 135, 1557S–1564S (2005).

5. Sanders, L. Branched-Chain Amino Acid Metabolism Disorders - Pediatrics. Merck Manuals (Professional) (2016).

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