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. 2019 Nov 22;10:5300. doi: 10.1038/s41467-019-13274-3

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 3 — Inactivation of MglA by MglB is regulated by a positive feedback loop. a Kinetics of GTP hydrolysis stimulated by MglB. Kinetics were monitored by fluorescence using a reagentless phosphate-binding assay at a fixed MglA concentration (1 µM) and increasing concentrations of MglB as indicated. b Time-dependent desensitization of MglA to MglB-stimulated GTP hydrolysis. MglA-GTP was incubated at 25 °C for increasing periods of time as indicated, before GTP hydrolysis was initiated by addition of MglB. c Desensitization of MglA from MglB is reversed by addition of MglB-sensitive MglA-GTP. MglB was added to MglB-insensitive (in orange) or MglB-sensitive (in blue) MglA-GTP and GTP hydrolysis was monitored for the duration needed for the MglB-sensitive sample to reach the plateau. Then, the same amount of MglB-sensitive MglA-GTP was added to both samples (arrow). Note that both kinetics curves reach the same plateau, indicating that all MglA-GTP has been converted to MglA-GDP in all experiments. d MglA-GDP, but not MglA-GTPγS, re-sensitizes MglA-GTP to MglB. MglA-GTP was incubated at 25 °C for 30 min, then MglB and 1 µM of either MglA-GDP or MglA-GTPγS was added.