Journal:Acta Cryst D:S2059798324001360

Characterization of novel mevalonate kinases from the tardigrade Ramazzottius varieornatus and the psychrophilic archaeon Methanococcoides burtonii

Lygie Esquirol, Janet Newman, Tom Nebl, Colin Scott, Claudia Vickers, Frank Sainsbury, Thomas S. Peat ^[1]

Molecular Tour
Many colors, odors and flavors found in nature are synthesized by the isoprenoid pathway. But besides pretty colors and smelly things, this pathway is essential for life, as it is also involved in the production of cholesterol, vitamins as well as in the production of secondary metabolites ranging from defensive compounds to antioxidants. It is found across the archaea, bacteria and eukaryote kingdoms. We all have the same pathway, and many enzymes are homologous across the three kingdoms of life, but of course they are also all a bit different. So we have a collection of enzymes performing the same reaction, but having different properties; some are highly inhibited by phosphorylated compounds, some like hot temperature, some like the cold to function, some are resistant to inhibition, etc. This diversity represents an incredible opportunity to further our understanding of the link between structure and function of enzymes.

Here we focused on the characterisation of two mevalonate kinases. Mevalonate kinase enzymes phosphorylate a mevalonate compound. This step in the pathway is a key control point for some bacteria and eukaryotes, where the mevalonate kinase activity is inhibited in presence of long phosphorylated compounds, made downstream of the pathway. This is called feedback-inhibition and it is one of the many mechanisms preventing the over-production of some of the potent secondary metabolites this pathway can produce. In archaea, however, the mevalonate kinases appear totally resistant to inhibition in presence of phosphorylated compounds and the pathway is controlled using other mechanisms.

We characterized two mevalonate kinases - one coming from a psychrophilic archaea, Methanococcoides burtonii (MKbur) and one coming from the tardigrade Ramazzottius varieornatus (MKvar). . The ‘N-term’ domains are in yellow and the ‘C-term’ domains in green. An MVA molecule is shown in cyan ball-and-sticks in the active site of MKvar and location of the GHMP Motifs I, II and III are indicated magenta, blue and grey respectively. (labelled helices are in red, loops in green and beta sheets in yellow). . The ‘N-term’ domains are in yellow and the ‘C-term’ domains in green. The GHMP Motifs I, II and III are indicated magenta, blue and grey respectively and the location of a Mg2+ ion involved in ATP binding is indicated by a cyan sphere. (labelled helices are in red, loops in green and beta sheets in yellow).

Zoom in around the active sites of MKbur (in orange) and MKvar (in green) are centered around the mevalonate:

• .
• .

The mevalonate is in cyan. Conserved amino acids are shown in ball-and-sticks, motifs I, II, III in magenta, blue and grey respectively.

Zoom in around the active sites of MKbur (in orange) and MKvar (in green) are centered around the ATP and the magnesium ion (Mg2+):

• .

The ATP is in shown in ball-and-sticks and the magnesium ion (Mg2+) represented as a cyan sphere (localization of ATP is estimated by alignment with rat mevalonate kinase (MKrat), crystallized with ATP in the active site (Fu et al., 2002^[2])). The loops involved in binding nucleotide moiety are marked in red and the lid is shown in yellow.

The tardigrade MKvar presents a classic feedback inhibition profile and is inhibited in presence of phosphorylated compounds such as farnesyl pyrophosphate (FPP) and geranyl pyrophosphate (GPP), whereas the MKbur is not. As expected MKbur is able to function at very low temperature as low as 4°C. The structures are resolved to 2 Å for MKvar and 2.2 Å for MKbur.

References

1. ↑ Esquirol L, Newman J, Nebl T, Scott C, Vickers C, Sainsbury F, Peat TS. Characterization of novel mevalonate kinases from the tardigrade Ramazzottius varieornatus and the psychrophilic archaeon Methanococcoides burtonii. Acta Crystallogr D Struct Biol. 2024 Mar 1;80(Pt 3):203-215. PMID:38411551 doi:10.1107/S2059798324001360
2. ↑ Fu Z, Wang M, Potter D, Miziorko HM, Kim JJ. The structure of a binary complex between a mammalian mevalonate kinase and ATP: insights into the reaction mechanism and human inherited disease. J Biol Chem. 2002 May 17;277(20):18134-42. Epub 2002 Feb 27. PMID:11877411 doi:10.1074/jbc.M200912200