# An atypical F-type ATPase is necessary for the function of the antibody cleavage system MIB-MIP in mycoplasmas

## TL;DR

Mycoplasmas use an unusual F-type ATPase to help evade the host's immune system by working with the MIB-MIP antibody cleavage system.

## Contribution

The study identifies a novel functional link between an atypical F-type ATPase and the MIB-MIP system in mycoplasmas for immune evasion.

## Key findings

- The atypical F1-like X0 ATPase is genetically and functionally linked to the MIB-MIP system in mycoplasmas.
- Mutant strains lacking functional ATPase are unable to resist antibody-mediated agglutination.
- The ATPase and MIB-MIP components co-purify, suggesting a physical interaction.

## Abstract

Mycoplasmas are genome-reduced, parasitic bacteria colonizing a broad range of organisms, including most livestock species and humans. They are often pathogenic, and have evolved an arsenal of mechanisms to counter their host’s immune response. Among these, the Mycoplasma Immunoglobulin Binding/Protease (MIB-MIP) system appears to be particularly important, and is conserved in the majority of mycoplasma species. Through genomics analysis, we show that MIB-MIP systematically co-occurs with a cluster of 7 coding sequences corresponding to an atypical Type 3 F-ATPase termed “F1-like X0”. Working in the model organism Mycoplasma mycoides subsp. capri, we first performed a proteomics analysis to confirm that this ATPase is indeed expressed. We then generated two mutant strains in which the putative ATPase was either fully deleted, or rendered catalytically inactive through replacement of a conserved lysine residue in the Walker A motif of the ATPase β-like subunit. Functional assays in presence of immune serum showed that both mutants are unable to protect themselves from agglutination by immunoglobulins despite the MIB-MIP system still being present. We then attempted to affinity-purify the atypical F1-like X0 ATPase from the membrane of its native host. Although the complex appears to be labile, under cross-linking conditions we were able to co-purify all its predicted components, as well as both MIB and MIP. These results allow us to attribute a function to Type 3 F-ATPase, namely to participate in the evasion from the humoral immune response in mycoplasmas, in conjunction with the MIB-MIP system through a currently unknown mechanism.

Mycoplasmas are small parasitic bacteria responsible for infections in a range of mammalian hosts, including humans and most livestock species. They take part in a constant arms race, evolving new mechanisms to try to evade or defeat their hosts’ complex immune systems. In this study, we describe an atypical F-ATPase known as Type 3, or F1-like X0, and show that it is genetically linked to MIB-MIP, as system dedicated to the capture and destruction of antibodies. Through phenotypic comparison of several mutant mycoplasma and co-purification attempts, we then show that this F-ATPase is indeed functionally linked to MIB-MIP, and even essential for the system to function. However, at this point the exact mechanism of interaction between all the partners is still unknown. Our study highlights the evolution of a complex molecular machine in mycoplasmas to combat the humoral immune response of their hosts. As the MIB-MIP- F1-like X0 ATPase system is conserved in mycoplasmas, it appears to be a promising target for the development of specific inhibitors, which could act as complement to classical antibiotics.