Susceptibility of Archaea to the Antibiotic Effect of the Parasporal Inclusion Proteins from Different Bacillus thuringiensis subspecies

The proteins of parasporal inclusions from three Bacillus thuringiensis subspecies (kurstaki, amagiensis, and monterrey) inhibited growth of methanogenic archaea of two species belonging to two genera, Methanobrevibacter arboriphilus and Methanosarcina barkeri. The minimal inhibitory concentrations of these proteins were 20 to 50 g/ml. Lysozyme exhibited similar bactericidal effect on archaea. The perspective of comparative studies on the effect of polyfunctional proteins on bacteria and archaea is discussed.     

Origin and evolution of the protein-repairing enzymes methionine sulphoxide reductases

Abstract The majority of extant life forms thrive in an O(2)-rich environment, which unavoidably induces the production of reactive oxygen species (ROS) during cellular activities. ROS readily oxidize methionine (Met) residues in proteins/peptides to form methionine sulphoxide [Met(O)] that can lead to impaired protein function. Two methionine sulphoxide reductases, MsrA and MsrB, catalyse the reduction of the S and R epimers, respectively, of Met(O) in proteins to Met. The Msr system has two known functions in protecting cells against oxidative damage. The first is to repair proteins that have lost activity due to Met oxidation and the second is to function as part of a scavenger system to remove ROS through the reversible oxidation/reduction of Met residues in proteins. Bacterial, plant and animal cells lacking MsrA are known to be more sensitive to oxidative stress. The Msr system is considered an important cellular defence mechanism to protect against oxidative stress and may be involved in ageing/senescence. MsrA is present in all known eukaryotes and eubacteria and a majority of archaea, reflecting its essential role in cellular life. MsrB is found in all eukaryotes and the majority of eubacteria and archaea but is absent in some eubacteria and archaea, which may imply a less important role of MsrB compared to MsrA. MsrA and MsrB share no sequence or structure homology, and therefore probably emerged as a result of independent evolutionary events. The fact that some archaea lack msr genes raises the question of how these archaea cope with oxidative damage to proteins and consequently of the significance of msr evolution in oxic eukaryotes dealing with oxidative stress. Our best hypothesis is that the presence of ROS-destroying enzymes such as peroxiredoxins and a lower dissolved O(2) concentration in those msr-lacking organisms grown at high temperatures might account for the successful survival of these organisms under oxidative stress.     

极端嗜热古菌的热休克蛋白

随着生物工程产业对于耐高温酶和菌体的需求, 极端嗜热古菌热休克蛋白(heat shock proteins, HSPs)的研究更受重视, 其热休克蛋白体系非常简洁, 不含HSP100s和HSP90s, 就是HSP70(DnaK)、HSP40、(DnaJ)和GrpE等嗜温古菌可能含有的在极端嗜热古菌中几乎不含有, 即仅包括HSP60, sHSP, prefoldin和AAA+蛋白四大类, 因此对其结构、功能和作用机制的研究在理论和实践上都特别有意义。系统地介绍了这四大类组分的结构、功能和作用机制和协同作用的研究进展, 论述了极端嗜热古菌热休克蛋白的系列研究难点和困惑, 展望了进一步的研究方向和重点。     

Discovery of two novel families of proteins that are proposed to interact with prokaryotic SMC proteins,and characterization of the Bacillus subtilis family members ScpA and ScpB

Structural maintenance of chromosomes (SMC) proteins are present in all eukaryotes and in many prokaryotes. Eukaryotic SMC proteins form complexes with various non-SMC subunits, which affect their function, whereas the prokaryotic homologues had no known non-SMC partners and were thought to act as simple homodimers. Here we describe two novel families of proteins, widespread in archaea and (Gram-positive) bacteria, which we denote 'segregation and condensation proteins' (Scps). ScpA genes are localized next to smc genes in nearly all SMC- containing archaea, suggesting that they belong to the same operon and are thus involved in a common process in the cell. The function of ScpA was studied in Bacillus subtilis, which also harbours a well characterized smc gene. Here we show that scpA mutants display characteristic phenotypes nearly identical to those of smc mutants, including temperature- sensitive growth, production of anucleate cells, formation of aberrant nucleoids, and chromosome splitting by the so-called guillotine effect. Thus, both SMC and ScpA are required for chromosome segregation and condensation. Interestingly, mutants of another B. subtilis gene, scpB, which is localized downstream from scpA, display the same phenotypes, which indicate that ScpB is also involved in these functions. ScpB is generally present in species that also encode ScpA. The physical interaction of ScpA and SMC was proven (i) by the use of the yeast two-hybrid system and (ii) by the isolation of a complex containing both proteins from cell extracts of B. subtilis. By extension, we speculate that interaction of orthologues of the two proteins is important for chromosome segregation in many archaea and bacteria, and propose that SMC proteins generally have non-SMC protein partners that affect their function not only in eukaryotes but also in prokaryotes.     

The COG database: new developments in phylogenetic classification of proteins from complete genomes

The database of Clusters of Orthologous Groups of proteins (COGs), which represents an attempt on a phylogenetic classification of the proteins encoded in complete genomes, currently consists of 2791 COGs including 45 350 proteins from 30 genomes of bacteria, archaea and the yeast Saccharomyces cerevisiae (http://www.ncbi.nlm.nih. gov/COG). In addition, a supplement to the COGs is available, in which proteins encoded in the genomes of two multicellular eukaryotes, the nematode Caenorhabditis elegans and the fruit fly Drosophila melanogaster, and shared with bacteria and/or archaea were included. The new features added to the COG database include information pages with structural and functional details on each COG and literature references, improvements of the COGNITOR program that is used to fit new proteins into the COGs, and classification of genomes and COGs constructed by using principal component analysis.     

Nucleotide bias causes a genomewide bias in the amino acid composition of proteins

We analyzed the nucleotide contents of several completely sequenced genomes, and we show that nucleotide bias can have a dramatic effect on the amino acid composition of the encoded proteins. By surveying the genes in 21 completely sequenced eubacterial and archaeal genomes, along with the entire Saccharomyces cerevisiae genome and two Plasmodium falciparum chromosomes, we show that biased DNA encodes biased proteins on a genomewide scale. The predicted bias affects virtually all genes within the genome, and it could be clearly seen even when we limited the analysis to sets of homologous gene sequences. Parallel patterns of compositional bias were found within the archaea and the eubacteria. We also found a positive correlation between the degree of amino acid bias and the magnitude of protein sequence divergence. We conclude that mutational bias can have a major effect on the molecular evolution of proteins. These results could have important implications for the interpretation of protein-based molecular phylogenies and for the inference of functional protein adaptation from comparative sequence data.     

PCNA activates the Holliday junction endonuclease Hjc

The resolving enzyme Hjc, which cleaves Holliday junctions with a high degree of structural specificity, is conserved in all archaea. Like RuvC in Escherichia coli, Hjc functions in the related processes of homologous recombination and double-strand break repair. In bacteria, the RuvAB complex binds Holliday junctions and catalyses ATP-dependent branch migration, but the equivalent proteins in archaea and eukarya are unknown. Here, we demonstrate that Hjc from Sulfolobus solfataricus forms a physical interaction with the sliding clamp PCNA via a C-terminal PCNA-interacting peptide (PIP) motif in Hjc. PCNA stimulates the Holliday junction cleavage activity of Hjc in vitro, and deletion of the PIP motif abrogates this effect. This is the first report of a functional interaction between a sliding clamp and a junction-resolving enzyme, and raises the possibility that PCNA could recruit a variety of different proteins to act on Holliday junctions in vivo.     

Identification of thermophilic species by the amino acid compositions deduced from their genomes

The global amino acid compositions as deduced from the complete genomic sequences of six thermophilic archaea, two thermophilic bacteria, 17 mesophilic bacteria and two eukaryotic species were analysed by hierarchical clustering and principal components analysis. Both methods showed an influence of several factors on amino acid composition. Although GC content has a dominant effect, thermophilic species can be identified by their global amino acid compositions alone. This study presents a careful statistical analysis of factors that affect amino acid composition and also yielded specific features of the average amino acid composition of thermophilic species. Moreover, we introduce the first example of a 'compositional tree' of species that takes into account not only homologous proteins, but also proteins unique to particular species. We expect this simple yet novel approach to be a useful additional tool for the study of phylogeny at the genome level.     

Identification of the new protein participating in the archaea motility regulation

A new family of archaeal proteins, CheM, having no homologues among bacteria and eukaryotes, was identified. Genes cheM are represented only in archaea possessing the chemotaxis and generally located close to che and fla loci. There is only one copy of the cheM gene in thermophilic and methanogenic archaea. Halophilic archaea have an additional paralog of the cheM gene. Mutant strains of Halobacterium salinarum R1 with deletions of the cheM1 (OE2402F) and cheM2 (OE2404R) genes were obtained. Mutant strains were not differ from the wild type strain by speed of movement in liquid medium but had appreciable differences in the diameter of a swarm on semi-liquid agar, indicative of reduced chemotaxis. It was demonstrated that the CheM2 protein from H. salinarum R1 co-isolates with protein CheY, the chemotaxis regulator in the conditions of its activation. The specific interaction between proteins CheM and CheY from hyperthermophilic archaea Pyrococcus horikoshii OT3 was also found. We suppose that CheM proteins provide adaptation of the chemotaxis system universal for bacteria and archaea to the specific archaeal flagellar motor apparatus.     

Why the Lipid Divide? Membrane Proteins as Drivers of the Split between the Lipids of the Three Domains of Life

Recent results from engineered and natural samples show that the starkly different lipids of archaea and bacteria can form stable hybrid membranes. But if the two types can mix, why don't they? That is, why do most bacteria and all eukaryotes have only typically bacterial lipids, and archaea archaeal lipids? It is suggested here that the reason may lie on the other main component of cellular membranes: membrane proteins, and their close adaptation to the lipids. Archaeal lipids in modern bacteria could suggest that the last universal common ancestor (LUCA) had both lipid types. However, this would imply a rather elaborate evolutionary scenario, while negating simpler alternatives. In light of widespread horizontal gene transfer across the prokaryotic domains, hybrid membranes reveal that the lipid divide did not just occur once at the divergence of archaea and bacteria from LUCA. Instead, it continues to occur actively to this day. Also see the video abstract here https://youtu.be/TdKjxoDAtsg .     

Influence of chromatin and single strand binding proteins on the activity of an archaeal MCM

The mini-chromosome maintenance (MCM) complex is the presumptive replicative helicase in archaea and eukaryotes. In archaea, the MCM is a homo-multimer, in eukaryotes a heterohexamer composed of six related subunits, MCM 2-7. Biochemical studies using naked DNA templates have revealed that archaeal MCMs and a sub-complex of eukaryotic MCM 4, 6 and 7 have 3' to 5' helicase activity. Here, we investigate the influence of the major chromatin proteins, Alba and Sul7d, of Sulfolobus solfataricus (Sso) on the ability of the MCM complex to melt partial duplex DNA substrates. In addition, we test the effect of Sso SSB on MCM activity. We reveal that Alba represents a formidable barrier to MCM activity and further demonstrate that acetylation of Alba alleviates repression of MCM activity.     

Protein secretion in the Archaea: multiple paths towards a unique cell surface

Archaea are similar to other prokaryotes in most aspects of cell structure but are unique with respect to the lipid composition of the cytoplasmic membrane and the structure of the cell surface. Membranes of archaea are composed of glycerol-ether lipids instead of glycerol-ester lipids and are based on isoprenoid side chains, whereas the cell walls are formed by surface-layer proteins. The unique cell surface of archaea requires distinct solutions to the problem of how proteins cross this barrier to be either secreted into the medium or assembled as appendages at the cell surface.     

Archaeal protein translocation crossing membranes in the third domain of life.

Proper cell function relies on correct protein localization. As a first step in the delivery of extracytoplasmic proteins to their ultimate destinations, the hydrophobic barrier presented by lipid-based membranes must be overcome. In contrast to the well-defined bacterial and eukaryotic protein translocation systems, little is known about how proteins cross the membranes of archaea, the third and most recently described domain of life. In bacteria and eukaryotes, protein translocation occurs at proteinaceous sites comprised of evolutionarily conserved core components acting in concert with other, domain-specific elements. Examination of available archaeal genomes as well as cloning of individual genes from other archaeal strains reveals the presence of homologues to selected elements of the bacterial or eukaryotic translocation machines. Archaeal genomic searches, however, also reveal an apparent absence of other, important components of these two systems. Archaeal translocation may therefore represent a hybrid of the bacterial and eukaryotic models yet may also rely on components or themes particular to this domain of life. Indeed, considering the unique chemical composition of the archaeal membrane as well as the extreme conditions in which archaea thrive, the involvement of archaeal-specific translocation elements could be expected. Thus, understanding archaeal protein translocation could reveal the universal nature of certain features of protein translocation which, in some cases, may not be readily obvious from current comparisons of bacterial and eukaryotic systems. Alternatively, elucidation of archaeal translocation could uncover facets of the translocation process either not yet identified in bacteria or eukaryotes, or which are unique to archaea. In the following, the current status of our understanding of protein translocation in archaea is reviewed.     

Stimulation of MCM helicase activity by a Cdc6 protein in the archaeon Thermoplasma acidophilum

Replicative DNA helicases are ring-shaped hexamers that play an essential role in chromosomal DNA replication. They unwind the two strands of the duplex DNA and provide the single-stranded (ss) DNA substrate for the polymerase. The minichromosome maintenance (MCM) proteins are thought to function as the replicative helicases in eukarya and archaea. The proteins of only a few archaeal organisms have been studied and revealed that although all have similar amino acid sequences and overall structures they differ in their biochemical properties. In this report the biochemical properties of the MCM protein from the archaeon Thermoplasma acidophilum is described. The enzyme has weak helicase activity on a substrate containing only a 3′-ssDNA overhang region and the protein requires a forked DNA structure for efficient helicase activity. It was also found that the helicase activity is stimulated by one of the two T.acidophilum Cdc6 homologues. This is an interesting observation as it is in sharp contrast to observations made with MCM and Cdc6 homologues from other archaea in which the helicase activity is inhibited when bound to Cdc6.     

Murein and pseudomurein cell wall binding domains of bacteria and archaea—a comparative view

The cell wall, a major barrier protecting cells from their environment, is an essential compartment of both bacteria and archaea. It protects the organism from internal turgor pressure and gives a defined shape to the cell. The cell wall serves also as an anchoring surface for various proteins and acts as an adhesion platform for bacteriophages. The walls of bacteria and archaea are mostly composed of murein and pseudomurein, respectively. Cell wall binding domains play a crucial role in the non-covalent attachment of proteins to cell walls. Here, we give an overview of the similarities and differences in the biochemical and functional properties of the two major murein and pseudomurein cell wall binding domains, i.e., the Lysin Motif (LysM) domain (Pfam PF01476) and the pseudomurein binding (PMB) domain (Pfam PF09373) of bacteria and archaea, respectively.     

Higher-level classification of the Archaea: evolution of methanogenesis and methanogens

We used a phylogenetic approach to analyze the evolution of methanogenesis and methanogens. We show that 23 vertically transmitted ribosomal proteins do not support the monophyly of methanogens, and propose instead that there are two distantly related groups of extant archaea that produce methane, which we have named Class I and Class II. Based on this finding, we subsequently investigated the uniqueness of the origin of methanogenesis by studying both the enzymes of methanogenesis and the proteins that synthesize its specific coenzymes. We conclude that hydrogenotrophic methanogenesis appeared only once during evolution. Genes involved in the seven central steps of the methanogenic reduction of carbon dioxide (CO(2)) are ubiquitous in methanogens and share a common history. This suggests that, although extant methanogens produce methane from various substrates (CO(2), formate, acetate, methylated C-1 compounds), these archaea have a core of conserved enzymes that have undergone little evolutionary change. Furthermore, this core of methanogenesis enzymes seems to originate (as a whole) from the last ancestor of all methanogens and does not appear to have been horizontally transmitted to other organisms or between members of Class I and Class II. The observation of a unique and ancestral form of methanogenesis suggests that it was preserved in two independent lineages, with some instances of specialization or added metabolic flexibility. It was likely lost in the Halobacteriales, Thermoplasmatales and Archaeoglobales. Given that fossil evidence for methanogenesis dates back 2.8 billion years, a unique origin of this process makes the methanogenic archaea a very ancient taxon.     

Compositional and Structural Features Related to Thermal Stability in the Archaea SRP19 and SRP54 Signal Recognition Particle Proteins

To survive at high temperature, thermophile organisms must adapt their biomolecules. In both nucleic acids and proteins, this adaptation involves a vast array of compositional and structural modifications. The archaea stand out as the only group of organisms that have species capable of growing at temperatures ranging from 0 to 110°C. In this study, we have used the archaea genome datasets to identify molecular trends related to thermal adaptation in the protein components (SRP19 and SRP54) of the signal recognition particle (SRP). Using comparative genomics and secondary structure homology modeling we have detected significant differences in the amino acids composition and distribution between the SRP proteins of thermophile and mesophile archaea. These include: a significant increase in the thermophile SRP proteins of the frequency of charged amino acids able to participate in electrostatic interactions which contribute to stabilize proteins; decreased content of both thermolabile and small/tiny amino acids which usually contribute to protein flexibility; and a significant increase in aliphatic and aromatic amino acids providing good covering and masking to produce hydrophobic pockets involved in stabilizing protein structure. Moreover, a detailed analysis of the four structural and functional domains of the SRP54 indicates a particularly robust correlation between the compositional properties of the M domain and the optimal growth temperature (OGT) of the archaea. The analysis of the bacterial SRP54(Ffh) shows similar adaptations to the OGT. Thus, natural selection has adapted the SRP proteins to the OGT of the archaea and bacteria species by modifying both, their amino acids composition and distribution.     

Functional annotation of class I lysyl-tRNA synthetase phylogeny indicates a limited role for gene transfer

Functional and comparative genomic studies have previously shown that the essential protein lysyl-tRNA synthetase (LysRS) exists in two unrelated forms. Most prokaryotes and all eukaryotes contain a class II LysRS, whereas most archaea and a few bacteria contain a less common class I LysRS. In bacteria the class I LysRS is only found in the alpha-proteobacteria and a scattering of other groups, including the spirochetes, while the class I protein is by far the most common form of LysRS in archaea. To investigate this unusual distribution we functionally annotated a representative phylogenetic sampling of LysRS proteins. Class I LysRS proteins from a variety of bacteria and archaea were characterized in vitro by their ability to recognize Escherichia coli tRNA(Lys) anticodon mutants. Class I LysRS proteins were found to fall into two distinct groups, those that preferentially recognize the third anticodon nucleotide of tRNA(Lys) (U36) and those that recognize both the second and third positions (U35 and U36). Strong recognition of U35 and U36 was confined to the pyrococcus-spirochete grouping within the archaeal branch of the class I LysRS phylogenetic tree, while U36 recognition was seen in other archaea and an example from the alpha-proteobacteria. Together with the corresponding phylogenetic relationships, these results suggest that despite its comparative rarity the distribution of class I LysRS conforms to the canonical archaeal-bacterial division. The only exception, suggested from both functional and phylogenetic data, appears to be the horizontal transfer of class I LysRS from a pyrococcal progenitor to a limited number of bacteria.