1.A.72. The Mercuric Ion Pore (Mer) Superfamily

The MerF protein encoded on plasmid pMER327/419 is an 81 residue polypeptide with two putative TMSs (Barkay et al., 2003).  It catalyzes uptake of Hg2+ in preparation for reduction by mercuric reductase. The MerF gene is found on mercury resistant plasmids from many Gram-negative bacteria, but the sequence of the protein from these plasmids is the same. Its two TMSs show limited sequence similarity with the first two TMSs of MerT (TC#1.A.72.3) and MerC (TC# 1.A.72.4). Some members of the MerF family have been designated MerH (Wilson et al., 2000).

The MerTP permeases catalyze uptake into bacterial cells of Hg2+  in preparation for its reduction by the MerA mercuric reductase. The Hgo produced by MerA is volatile and passively diffuses out of the cell. The merT and merP genes are found on mercury resistance plasmids and transposons of Gram-negative and Gram-positive bacteria but are also chromosomally encoded in some bacteria. MerT consists of about 130 amino acids and has 3 transmembrane helical segments. Evidence for direct interactions between the cytoplasmic face of MerT and theN-terminus of MerA have been presented (Schué et al. 2009). Operon analyses have been reported by Barkay et al., 2003Miller,1999Velasco et al., 1999

MerP is a periplasmic Hg2+ -binding receptor of about 70-80 amino acids, synthesized with a cleavable N-terminal leader. It is homologous to the N-terminal heavy metal binding domains of the Copper-and cadmium-transporting P-type ATPases. The 3-D structure of MerP from Ralstonia metallidurans has been solved to 2 Å resolution (Serre et al., 2004Qian et al.,1998). It is 91 aas long with its leader sequence, is monomeric, and binds a single Hg2+ion. Hg2+  is bound to a sequence GMTCXXC found in metallochaperones as well as metal-transporting ATPases. The fold is βαββαβ, called the ''ferridoxin-like fold''.

MerT homologues have been identified in which the 3 TMS MerT is fused to a MerP ''heavy metal associated'' (HMA) domain possibly via a linker region that includes a fourth TMS (see 1.A.72.3.3). HMA domains of ~30 aas are found in MerP, copper chaperone proteins, mercuric reductase, and at the N-termini of both copper and heavy metal P-type ATPases, sometimes in multiple copies.

The MerC protein encoded on the IncJ plasmid pMERPH of the Shewanella putrefaciens mercuric resistance operon is 137 amino acids in length and possesses four putative transmembrane α-helical spanners (TMSs). It has been shown to bind and take up Hg2+ ions. merC genes are encoded on several plasmids of Gram-negative bacteria and may also be chromosomally encoded. MerC proteins are homologous to other bacterial Hg2+ bacterial transporters ( Inoue et al., 1990; Peters et al., 1991; Mok et al., 2011; Yamaguchi et al., 2007).

The merE gene of transposon Tn21, a pE4 plasmid that contained the merR gene of plasmid pMR26 from Pseudomonas strain K-62, and the merE gene of Tn21 from the Shigella flexneri plasmid NR1 (R100) conferred hypersensitivity to CH3Hg2+  and Hg2+, taking up significantly more CH3Hg2+ and Hg2+ than the isogenic strain (Kiyono et al. 2009). The MerE protein encoded by pE4 was localized in the membrane cell fraction, but not in the soluble fraction. Kiyono et al. (2009) suggested that the merE gene is a broad mercury transporter mediating the transport of both CH3Hg2+  and Hg2+  across the bacterial membrane.

The common origin of all Mer superfamily members has been established (Mok et al., 2011).  The common elements are included in TMSs 1-2.

The transport reaction catalyzed by Mer Superfamily members is:

Hg2+ or methyl-Hg2+ (out) →  Hg2+ or methyl-Hg2+ (in)



This family belongs to the Mercuric Ion Pore (Mer) Superfamily.

 

References:

Yamaguchi, A., Tamang D., and Saier M. (2007). Mercury Transport in Bacteria. [DOI: 10.1007/s11270-007-9334-z]



Amin, A., A. Sarwar, M.A. Saleem, Z. Latif, and S. Opella. (2019). Expression and Purification of Transmembrane Protein MerE from Mercury-Resistant. J Microbiol Biotechnol 29: 274-282.

Barkay, T., S.M. Miller, and A.O. Summers. (2003). Bacterial mercury resistance from atoms to ecosystems. FEMS Microbiol. Rev. 27: 355-384.

Howell, S.C., M.F. Mesleh, and S.J. Opella. (2005). NMR structure determination of a membrane protein with two transmembrane helices in micelles: MerF of the bacterial mercury detoxification system. Biochemistry 44: 5196-5206.

Hwang, H., A. Hazel, P. Lian, J.C. Smith, J.C. Gumbart, and J.M. Parks. (2019). A Minimal Membrane Metal Transport System: Dynamics and Energetics of mer Proteins. J Comput Chem. [Epub: Ahead of Print]

Inoue, C., K. Sugawara, and T. Kusano. (1990). Thiobacillus ferrooxidans mer operon: sequence analysis of the promoter and adjacent genes. Gene 96: 115-120.

Kiyono, M., Y. Sone, R. Nakamura, H. Pan-Hou, and K. Sakabe. (2009). The MerE protein encoded by transposon Tn21 is a broad mercury transporter in Escherichia coli. FEBS Lett. 583: 1127-1131.

Kusano, T., G.Y. Ji, C. Inoue, and S. Silver. (1990). Constitutive synthesis of a transport function encoded by the Thiobacillus ferrooxidans merC gene cloned in Escherichia coli. J. Bacteriol. 172: 2688-2692.

Miller, S.M. (1999). Bacterial detoxification of Hg(II) and organomercurials. Essays Biochem 34: 17-30.

Morby, A.P., J.L. Hobman, and N.L. Brown. (1995). The role of cysteine residues in the transport of mercuric ions by the Tn501 MerT and MerP mercury-resistance proteins. Mol. Microbiol. 17: 25-35.

Peters, S.E., J.L. Hobman, P. Strike, and D.A. Ritchie. (1991). Novel mercury resistance determinants carried by IncJ plasmids pMERPH and R391. Mol. Gen. Genet. 228: 294-299.

Qian, H., L. Sahlman, P.O. Eriksson, C. Hambraeus, U. Edlund, and I. Sethson. (1998). NMR solution structure of the oxidized form of MerP, a mercuric ion binding protein involved in bacterial mercuric ion resistance. Biochemistry 37: 9316-9322.

Sasaki, Y., T. Minakawa, A. Miyazaki, S. Silver, and T. Kusano. (2005). Functional dissection of a mercuric ion transporter, MerC, from Acidithiobacillus ferrooxidans. Biosci. Biotechnol. Biochem. 69: 1394-1402.

Schué, M., L.G. Dover, G.S. Besra, J. Parkhill, and N.L. Brown. (2009). Sequence and analysis of a plasmid-encoded mercury resistance operon from Mycobacterium marinum identifies MerH, a new mercuric ion transporter. J. Bacteriol. 191: 439-444.

Serre, L., E. Rossy, E. Pebay-Peyroula, C. Cohen-Addad, and J. Covès. (2004). Crystal structure of the oxidized form of the periplasmic mercury-binding protein MerP from Ralstonia metallidurans CH34. J. Mol. Biol. 339: 161-171.

Sugio, T., T. Komoda, Y. Okazaki, Y. Takeda, S. Nakamura, and F. Takeuchi. (2010). Volatilization of metal mercury from Organomercurials by highly mercury-resistant Acidithiobacillus ferrooxidans MON-1. Biosci. Biotechnol. Biochem. 74: 1007-1012.

Velasco, A., P. Acebo, N. Flores, and J. Perera. (1999). The mer operon of the acidophilic bacterium Thiobacillus T3.2 diverges from its Thiobacillus ferrooxidans counterpart. Extremophiles 3: 35-43.

Venturi, E., K. Mio, M. Nishi, T. Ogura, T. Moriya, S.J. Pitt, K. Okuda, S. Kakizawa, R. Sitsapesan, C. Sato, and H. Takeshima. (2011). Mitsugumin 23 forms a massive bowl-shaped assembly and cation-conducting channel. Biochemistry 50: 2623-2632.

Wilson, J.R., C. Leang, A.P. Morby, J.L. Hobman, and N.L. Brown. (2000). MerF is a mercury transport protein: different structures but a common mechanism for mercuric ion transporters? FEBS Lett. 472: 78-82.



1.A.72.1 The MerF Mercuric Ion (Hg²⁺) Uptake (MerF) Family


Examples:

TC#NameOrganismal TypeExample
1.A.72.1.1
The MerF mercuric ion uptake transporter of 81 aas and 2 TMSs. The NMR structure of the helix-loop-helix core domain of MerF has been determined with a backbone RMSD of 0.58 Å (Howell et al. 2005). Moreover, the fold of this polypeptide demonstrates that the two vicinal pairs of cysteine residues, shown to be involved in the transport of Hg++ across the membrane, are exposed to the cytoplasm. This finding differs from earlier structural and mechanistic models that were based primarily on the somewhat atypical hydropathy plot for MerF and related transport proteins (Howell et al. 2005). The apo state positions one of the cysteine pairs closer to the periplasmic side of the membrane, while in the bound state, the same pair approaches the cytoplasmic side (Hwang et al. 2019). This is consistent with the functional requirement of accepting Hg2+ from the periplasmic space, sequestering it on acceptance, and transferring it to the cytoplasm. Conformational changes in the TMSs facilitate the functional interaction of the two cysteine pairs. Free-energy calculations provide a barrier of 16 kcal/mol for the association of the periplasmic Hg2+-bound protein, MerP, with MerF, and 7 kcal/mol for the subsequent association of MerF's two cysteine pairs (Hwang et al. 2019).

Bacteria

MerF of plasmid pMER327/419 of Pseudomonas aeruginosa

 
1.A.72.1.2

Heavy metal transporter

Bacteria

HM transporter of Arcobacter butzleri (A8EUY8)

 
1.A.72.1.3

MerT (97aas)/MerP (93aas) (in a single operon with a transglutaminase (COG1305)).

γ-Proteobacteria

MerTP of Haemophilus influenzae
MerT (Q57347)
MerP (P71365)

 


1.A.72.2 The MerH Mercuric Ion (Hg²⁺) Permease (MerH) Family


Examples:

TC#NameOrganismal TypeExample
1.A.72.2.1

Hg2+ transporter, MerH (171aas; 4 TMSs) (transports mercuric ions via a pair of essential cysteine residues, but only when coexpressed with the mercuric reductase) (Schué et al., 2009).

Actinobacteria

MerH of Mycobacterium marinum (B2I419)

 
1.A.72.2.2

MerC homologue (129aas; 4 TMSs)

MerC homologue of Gemmatimonas aurantiaca

 


1.A.72.3 The MerTP Mercuric Ion (Hg²⁺) Permease (MerTP) Family


Examples:

TC#NameOrganismal TypeExample
1.A.72.3.1

MerT/P

Bacteria

MerT/P of Ralstonia eutropha (Q6UP69)

 
1.A.72.3.2

Putative MerT-MerP fusion protein of 200 aas (3 TMSs)

Bacteroidetes

 MerT-MerP of Chryseobacterium gleum (C0YI47)

 
1.A.72.3.3

Putative MerT-MerP fusion protein of 199 aas (3-4 TMSs)

Verrucomicrobia

MerT-MerP of Methylacidiphilum infernorum (B3DYY6)

 
1.A.72.3.4

Putative MerT-MerP fusion protein of 196 aas (3 TMSs)

Bacteroidetes

MerT-MerP of Spirosoma linguale (D2QV66)

 
1.A.72.3.5

Mercuric ion uptake system, MerT-P/MerP

Bacteria

MerT-P/MerP of Tenacibaculum discolor
MerT-P (204aas; H6WCN3)
MerP (113aas; H6WCN4) 

 
1.A.72.3.6

Mercury transporter, MerT, of 129 aas and 3 TMSs.

MerT of Histidinibacterium lentulum

 


1.a.72.4 The MerC Mercuric Ion (Hg²⁺) Permease (MerC) Family


Examples:

TC#NameOrganismal TypeExample
1.A.72.4.1

MerC

Bacteria

MerC of the IncJ plasmid pMERPH of Shewanella putrefaciens

 
1.A.72.4.2

Mercuric transport channel protein, MerC, of 144 aas and 4 TMSs.  Cys-23 and Cys-26 of the protein were involved in Hg2+-recognition/uptake, but Cys-132 and Cys-137 were not (Sasaki et al. 2005). E. coli cells producing MerC were hypersensitive to CdCl2. In this case, mutation of His72 rendered the host cells less CdCl2 sensitive, whereas none of the Cys residues affected it. E. coli cells expressing a merC-deletion mutant, in which the coding-sequence of the carboxy-terminal cytoplasmic region was removed, retained Hg2+ hypersensitivity and showed about 55% HgCl2 uptake ability compared to that of the one expressing the intact merC, indicating that this region is not essential for Hg2+ uptake. Coexpression of the A. ferrooxidans gene encoding mercuric reductase (merA) and the merC deletion mutation conferred HgCl2 tolerance to E. coli host cells. Under this condition, the merC deletion gene product was exclusively present as a monomer (Sasaki et al. 2005).

MerC of Acidithiobacillus ferrooxidans (Thiobacillus ferrooxidans)

 


1.a.72.5 The MerE Mercuric Ion (Hg²⁺) Permease (MerE) Family


Examples:

TC#NameOrganismal TypeExample
1.A.72.5.1

The Mercuric ion (Hg2+) uptake transporter, MerE (78aas; 2 TMSs).

Bacteria

MerE of transposon Tn21 of E. coli (Q57069)

 
1.A.72.5.2

MerE mercury resistance protein of 89 aas and 2 TMSs.  It has been purified and characterized, and has proven useful for bioremediation (Amin et al. 2019).

MerE of Bacillus cereus