PhoH2 proteins couple RNA helicase and RNAse activities - PubMed (original) (raw)

Review

. 2020 Apr;29(4):883-892.

doi: 10.1002/pro.3814. Epub 2020 Jan 7.

Affiliations

Review

PhoH2 proteins couple RNA helicase and RNAse activities

Emma S V Andrews et al. Protein Sci. 2020 Apr.

Abstract

PhoH2 proteins are found in a very diverse range of microorganisms that span bacteria and archaea. These proteins are composed of two domains: an N-terminal PIN-domain fused with a C-terminal PhoH domain. Collectively this fusion functions as an RNA helicase and ribonuclease. In other genomic contexts, PINdomains and PhoHdomains are separate but adjacent suggesting association to achieve similar function. Exclusively among the mycobacteria, PhoH2 proteins are encoded in the genome with an upstream gene, phoAT, which is thought to play the role of an antitoxin (in place of the traditional VapB antitoxin that lies upstream of the 47 other PINdomains in the mycobacterial genome). This review examines PhoH2 proteins as a whole and describes the bioinformatics, biochemical, structural, and biological properties of the two domains that make up PhoH2: PIN and PhoH. We review the transcriptional regulators of phoH2 from two mycobacterial species and speculate on the function of PhoH2 proteins in the context of a Type II toxin-antitoxin system which are thought to play a role in the stress response in bacteria.

Keywords: AAA+ helicase; PIN-PhoH; PIN-domain; PhoH domain; PhoH2; RNAse; ribonuclease; toxin-antitoxin.

© 2019 The Protein Society.

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Figures

Figure 1

Figure 1

Domain organization, schematic of phoATphoH2 and phylogeny of PhoH2 proteins. (a) PhoH2 proteins are composed of a fusion between a PIN‐domain and a PhoHdomain separated by a flexible linker region. (b) phoAT overlaps the start of phoH2 similar to vapB of vapBC toxin–antitoxin systems. (c) Schematic of universal tree4 indicating phylum in which Ylak, COG1875 proteins can be found (red boxes). Last universal common ancestor (LUCA) and circular nodes correspond to last common ancestors

Figure 2

Figure 2

Conservation of PhoAT among closely related mycobacteria. (a) PhoAT sequence from members of the

M. tuberculosis

complex. (b) PhoAT sequence from more distantly related mycobacteria. Image was created using Geneious version 7.1 (Biomatters)

Figure 3

Figure 3

Hidden Markov model that defines PIN‐domain proteins. Based on a multiple sequence alignment the height of each letter defines the significance of the amino acid at that position within the PIN‐domain family. The width of each letter defines the significance of this position in the identification of the PIN‐domain family. The dark and light pink regions are where insertions have occurred. The three conserved amino acids (D, E, D) at Positions 4, 40, and 93 characterize the PIN‐domain protein family along with polar residues located after the first conserved aspartic acid as shown by asterisks. Generated using Pfam1

Figure 4

Figure 4

VapC PIN‐domain dimer and tetramer. (a) Ribbon diagram of VapCPAE2754 dimer. The three conserved acidic residues are labeled shown as sticks. (b) Ribbon diagram of VapCPAE2754 dimer rotated 90°. (c) Electrostatic surface diagram of PAE2754 tetramer. Negative (red) and Positive (blue) charges show where ssRNA may bind to positively charged residues at the entrance of the tunnel. Images were made in PyMOL V1.3 using PDB IV8P

Figure 5

Figure 5

Hidden Markov model that defines PhoHdomain proteins. Based on a multiple sequence alignment, the height of each letter defines the significance of the amino acid at that position within the PhoH‐domain family. The width of each letter defines the significance of this position in the identification of the PhoH‐domain family. The dark and light pink regions are where insertions have occurred. Conserved regions are comparable to motifs for ATPase activity found across each of the helicase families and two motifs unique to PhoH involved with nucleic interactions (RRB 1 and 2). Generated using Pfam1

Figure 6

Figure 6

Structures of the PhoHdomain of PhoH2 from C. glutamicum. (a) Active hexameric conformation of PhoH. (b) Electrostatic map of PhoH hexamer (blue positive charge and red negative charge) showing the positive charges present at the predicted entrance of the tunnel. (c) Magnified view of the PhoH‐domain active site showing each of the conserved residues and motifs labeled and colored as positions from HMM (Q, WA, RRB1, RRB2, WB, SI, SRH, Motif III, and SII). Sulfate ions were identified in the structure (shown as yellow and orange sticks) and show the position where the γ and β phosphate of ATP binds in the active site. Motifs Q, W A/B, RRB1, Motif III, SI, and SII interact with RRB2 and the SRH from the neighboring monomer. Images were made in PyMol V 1.3 using PDB 3B85

Figure 7

Figure 7

Genomic context of phoH2 genes from M. tuberculosis and M. smegmatis. Genomic context of phoH2 genes from (a) M. tuberculosis and (b) M. smegmatis. phoAT is indicated by the short black arrow directly upstream of phoH2. Rv1096 encodes a glycosyl hydrolase, Rv1097c a putative glycine and proline rich membrane protein and MS_5250 encodes a hypothetical protein of unknown function, CalR9, MS_5246 encodes a nitroreductase and MS_5245 a universal stress protein. Adapted from Xbase (

http://xbase.ac.uk

) and BLAST (

http://blast.ncbi.nlm.nih.gov/Blast.cgi

)

Figure 8

Figure 8

Genomic positions of predicted transcription factor binding sites. Arrows indicate the position of the five predicted transcription factor binding sites. phoAT is indicated by the black block arrow upstream of phoH2. Adapted from the Tuberculosis Database (TBDB;

http://www.tbdb.org/

)

Figure 9

Figure 9

Model of the proposed phoH2 regulation and PhoH2 activity

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