Calculating metalation in cells reveals CobW acquires CoII for vitamin B12 biosynthesis while related proteins prefer ZnII

Tessa R. Young*, Maria Alessandra Martini, Andrew W. Foster, Arthur Glasfeld, Deenah Osman, Richard J Morton, Evelyne Deery, Martin J. Warren, Nigel J. Robinson*

*Corresponding author for this work

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Abstract

Protein metal-occupancy (metalation) in vivo has been elusive. To address this challenge, the available free energies of metals have recently been determined from the responses of metal sensors. Here, we use these free energy values to develop a metalation-calculator which accounts for inter-metal competition and changing metal-availabilities inside cells. We use the calculator to understand the function and mechanism of GTPase CobW, a predicted Co -chaperone for vitamin B . Upon binding nucleotide (GTP) and Mg , CobW assembles a high-affinity site that can obtain Co or Zn from the intracellular milieu. In idealised cells with sensors at the mid-points of their responses, competition within the cytosol enables Co to outcompete Zn for binding CobW. Thus, Co is the cognate metal. However, after growth in different [Co ], Co -occupancy ranges from 10 to 97% which matches CobW-dependent B synthesis. The calculator also reveals that related GTPases with comparable Zn affinities to CobW, preferentially acquire Zn due to their relatively weaker Co affinities. The calculator is made available here for use with other proteins.
Original languageEnglish
Article number1195
Number of pages15
JournalNature Communications
Volume12
Issue number1
Early online date19 Feb 2021
DOIs
Publication statusE-pub ahead of print - 19 Feb 2021

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