Magnetite biomineralization in Magnetospirillum magneticum is regulated by a switch-like behavior in the HtrA protease MamE

Abstract

Magnetotactic bacteria are aquatic organisms that produce subcellular magnetic particles in order to orient in the earth's geomagnetic field. MamE, a predicted HtrA protease required to produce magnetite crystals in the magnetotactic bacterium Magnetospirillum magneticum AMB-1, was recently shown to promote the proteolytic processing of itself and two other biomineralization factors in vivo. Here, we have analyzed the in vivo processing patterns of three proteolytic targets and used this information to reconstitute proteolysis with a purified form of MamE. MamE cleaves a custom peptide substrate with positive cooperativity, and its autoproteolysis can be stimulated with exogenous substrates or peptides that bind to either of its PDZ domains. A misregulated form of the protease that circumvents specific genetic requirements for proteolysis causes biomineralization defects, showing that proper regulation of its activity is required during magnetite biosynthesis in vivo. Our results represent the first reconstitution of the proteolytic activity of MamE and show that its behavior is consistent with the previously proposed checkpoint model for biomineralization.Magnetotactic bacteria assemble iron-based magnetic crystals called magnetosomes into chains within their cells, allowing them to passively align in and navigate along magnetic fields (1, 2). Understanding the mechanism of biomineralization in these organisms can provide novel strategies for manipulating transition metal-based nanomaterials in vitro (3-5). Genetic analyses have identified a set of genes, called biomineralization factors, whose deletions disrupt or eliminate magnetite crystal formation (6 -10). Two of these, mamE and mamO, encode predicted trypsin-like proteases required to produce magnetite in the model magnetotactic organism, Magnetospirillum magneticum AMB-1 (6). Disrupting either gene abolishes the formation of magnetite crystals without disturbing the production of their surrounding membrane compartment, showing that each protein is required for initiating magnetite biosynthesis (6,11,12).Cells with a catalytically inactive (E PD ) allele of mamE show an intermediate biomineralization phenotype in which they produce small magnetite particles. Wild-type AMB-1 has a distribution of crystal sizes centered at 50 -60 nm in diameter, but the size distribution in the E PD cells is centered at ϳ20 nm. Interestingly, ϳ97% of the crystals in the E PD strain are smaller than 35 nm, the point above which magnetite particles become paramagnetic and can hold a stable magnetic dipole. The correlation between mineral sizes in the E PD strain and the superparamagnetic to paramagnetic transition point lead to a model predicting that cells produce small superparamagnetic crystals until an unknown signal activates MamE, promoting maturation to paramagnetic particles (13).MamE is a member of the HtrA/DegP family of trypsin-like proteases, a ubiquitous family of enzymes that controls various aspects of protein quality control (14). The family is characterized b...

Large‐scale survey and database of high affinity ligands for peptide recognition modules

“…Nevertheless, our database should prove useful to help the generation of new annotations for future functional ligands with optimized affinities and to guide further exploration of PRM families that have not yet been studied. Moreover, the database can be used to predict natural interaction partners and provide insights into potential cellular functions of PRMs, as has been shown previously by us and others (Schon et al, 2002;Slivka et al, 2009;Zhang et al, 2009;Reimand et al, 2012;Kelil et al, 2016;Hershey et al, 2016). YWHAE (14-3-3) YWHAZ (14-3-3) YWHAG (14-3-3) A-C Depicted are the 17 PRMs for which the ligand in the structure contains (A) pSer/pThr, (B) pTyr, or (C) meArg/meLys.…”

“…These studies have provided 7,063 unique and validated peptide binders for a total of 342 domains, and the resulting specificity maps have revealed the versatile and specific nature of these modules and their interactions. Moreover, the database of optimal ligands for PDZ, SH3 and WW domains has proven to be highly useful for predicting and validating natural interactions, developing intracellular inhibitors of natural interactions, and guiding structural studies to better understand the details of molecular recognition (Schon et al , 2002; Slivka et al , 2009; Zhang et al , 2009; Kelil et al , 2016; Hershey et al , 2016).…”

WITHDRAWN: OxyR-Like Improve Cell's Hydrogen Peroxide Tolerance via Participate in Monocyte Chemotaxis and Oxidative Phosphorylation Regulation in Magnetospirillum Gryphiswaldense MSR-1

“…mamJ, mamK and mamY help the single magnetosome arranging into chains [15][16][17]. mamE and mamO are essential during magnetic crystal formation [18,19]. mms6 and mms7 genes take part in the morphology control of nanoparticles [20][21][22].…”

A protease-mediated switch regulates the growth of magnetosome organelles in Magnetospirillum magneticum

“…Here, we find that the catalytic activity of MamE plays a central role in the progression of magnetosome membrane growth. MamE proteolytically processes itself and two other biomineralization factors, MamO and MamP (21). MamO is required for MamE protease activation (22), and we find it acts as an upstream regulator of MamE for magnetosome membrane growth.…”