Function
Two enzymes are responsible for the oxalate that is produced in the Burkholderia species. The first enzyme, ObcA, catalyzes the formation of a tetrahedral C6-CoA adduct from the substrates acetyl-COA and oxaloacetate. The second enzyme, ObcB, produces three products from the C6-CoA adduct. These products are oxalate, acetoacetate and CoA. The oxalate produced from the , is necessary for bacterial growth and maintaining environmental pH [1].
Disease
Species of Burkholderia can be involved in plant or human pathogenesis. Several diseases, such as B. Glumae which causes bacterial panicle blight in rice [2], or B. cepacia, which is an opportunistic pathogen in immunocompromised individuals, like those with cystic fibrosis or chronic granulomatous disease [3]. It can also be involved with B. pseudomallei which can cause meliondosis, a lethal infection that leads to the formation of abscesses in internal organs [4].
Relevance
By studying the two mono-functional enzymes, ObcA and ObcB, there is a better understanding of the underlying molecular basis. This bifunctional enzyme, Obc 1, can be used here for oxalogenesis. Researching and finding out more about these enzymes can help advance knowledge and potentially develop ways to control diseases associated with the Burkholderia species [1].
Structural highlights
The of this protein is made up of mostly alpha helices (pink), with some additional beta sheets (yellow).
The of this protein has two domains, the N-domain (navy) and the C-Domain (gray). The two domains in this protein, Obc 1, mediate oxalogenesis. The N-domain consists of an Obc B activity-exhibiting C-terminal region (Arg-529 to Gln-1106). It was found that there were no extensive interactions between the two domains, and researchers decided to focus on the C-domain. The C-domain (Arg-530 to Gln-1106) has features common to canonical alpha/beta hydrolyses [1]. When looking at the it is hard to distinguish between the different parts of the protein. The red balls in this view represent oxygen molecules. This view can give a better insight on the size, shape, and representation of the complete molecule. The protein seems to be hydrophobic and hydrophilic. The (C3 H8 O3) shown is glycerol, which is bound to the Obc 1 C-domain [1]. Arg-935, His-1069, and Asp-997 are the residues that make up the . The catalytic triad is located in the C-domain and is crucial to oxalate production. A change in the catalytic triad would most likely result in loss of function of the protein [1].
The of the C-domain in a crevice between the cap domain and the alpha/beta hydrolase fold, and the position of the catalytic Ser-935.
A loop of Ser-785–Thr-786 –Pro-787 and two arginine residues (Arg-856 and Arg-999) are in the active site. Arg-856 and Arg-999, were found to be necessary
for the activity of Obc 1, showing that the two residues could serve as an oxyanion binding site in Obc1. The C-domain consists of . The first region (navy), Ser-740 to Gln-1106, and forms an alpha/beta hydrolase fold. The second subdomain (red), Arg-529 to Ala-739, is located over a concave region formed by an alpha/beta hydrolase fold, resulting in a crevice between the two regions. The second domain is referred to as the cap domain [1].
The was found to be important for maintaining the structural integrity of a Ser-785–Thr-786 –Pro-787 loop near catalytic Ser-935 [1]. The catalytic triad is located in the loop region, and these residues are clustered in a in the C-domain, and their relative locations are conserved in other alpha/beta hydrolase [5].
For kinetic conditions, C6-CoA adduct was found to be produced from ObcA. It was stable and could not be converted into CoA in the absence of Obc1, meaning the formation of CoA from the adduct is enzyme-dependent. The activity of Obc1 was measured in two different ways, both by the production of different products. In both experiments, the reaction mixture contained Co2+ ion as the most effective ion for ObcA activity [1].
