Metals from cell to environment: Connecting Metallomics with other omics

Research activities and data collection of metals present in living organisms are called as “metallomics”. In metallomics, biomolecules incorporating metal ions viz. metalloenzymes and metalloproteins, are known as “metallomes”. Metallomics aims to identify metallomes of living organisms and to annotate the physiological signifi cance as well as the biological functions. However, in order to ascertain metallomics to be the part of biometal science, recent analytical technologies like chemical speciation are required to analyze the metallomes. Environmental applications like bioleaching, phytoremediation of soil by using microbes, and to deal with the uptake, transport, storage of trace metals necessary for protein functions and biomarkers identifi cation under ecotoxicological studies really require metallomics involvement. As an interdisciplinary research area, metallomics cover plant and animal physiology, nutrition and become a potential candidate in pharmacology, biogeochemistry and clinical chemistry. Metallomics uses analytical and spectroscopic methods to fi nd the quantitative and qualitative information about metal ions that are present as ligands a well multifaceted biological matrix in trace amounts or occur as noncovalent complexes in order to perform different biological processes. Latest spectroscopic methods along with in-silico approaches including bioinformatics are the important tools needed for research activities in metallomics. The present review highlights the basics of metallomics in biological sciences and its emergence as a novel omics era in relation to other fi elds. Besides the above aspects, applications and future prospects of metallomics have been highlighted. Review Article Metals from cell to environment: Connecting Metallomics with other omics Vijeta Singh1*and Kusum Verma2 1Amity Institute of Biotechnology, Amity University, Gautam Budha Nagar, India 2Department of Microbiology, KNIPSS, Sultanpur-228001, U.P., India Received: 03 February, 2018 Accepted: 10 March, 2018 Published: 12 March, 2018 *Corresponding author: Vijeta Singh, Amity Institute of Biotechnology, Amity University, Gautam Budha Nagar, India, Tel No: +91-9990250103; E-mail:


Introduction from Metals to Omics
Biological systems utilize metal ions for their fundamental processes like signaling, catalysis and gene expression. Few metals due to their metallic properties cause various diseases like Pt, Cr, as are carcinogenic whereas Pt, Co, Cr, Ni, Au cause immunological diseases. Mercury is teratogenic and causes diseases related to embryos. Nephrological disorders are caused by Cd and U whereas Al, Hg, and Mn are neurotoxic [1]. The essential transition metals e.g. Zn, Fe, Se, Cu are generally used by cells as cofactors of enzymes, can also potentially catalyze cytotoxic reactions [2]. Further, metals ions govern important properties like the folding of proteins, their conformation, assembly, stability, and catalysis. Many metals mostly transition metals (Mo, Fe, Zn, Cu, Mn) are required as a cofactor of various enzymes for their specifi c activities [3]. Metal ions also up-and down-regulate protein expression in the cells. Metallothionein proteins play a crucial role in cellular homeostasis and detoxifi cation responses [4]. Therefore, metal ion concentration in the cell, their integration in metalloproteins and their allocation to different cell organelles has been considered to be under tight regulation [5]. The DNA, RNA, and biosynthesis of protein require various metalloenzymes and related metal ions [6]. In this line, nowadays with the help of the high-throughput approaches the complete genetic blueprint of various organisms has been studied which resulted in various ''omics''; the disciplines aiming at the annotation of a particular class of components of a living organism in its ensemble [7]. Therefore, metallomics is a very needful area to be focused scientifi cally among researchers along with other omics such as genomics and proteomics.
It is very important to defi ne the terms and render their use systematically [10]. Among various omics, metallomics is defi ned to apprehend the metal-dependent life processes [11], or the assemblage of metals related research in the living organisms. However, the word "metallome" was fi rst given by Williams who referred to it as an element distribution, metal ion concentration, or as free elements present in the cellular organelles, cell or organism [12]. Under metallomics, ''metallomes'' is a general annotation for metalloenzymes, DOI: http://dx.doi.org/10.17352/ojps.000008 metalloproteins, and many other biomolecules containing metal ions which are similar to genomes for genomics (the study of nucleic acids) and proteomes for proteomics (the study of proteins and amino acids). The main research objectives of metallomics involve the study of physiological and biological functions of living systems and identifi cation of metallomes, however, chemical speciation, the important technology required for the establishment of metallomics is an integral branch of biometal science [13]. Now metallomics has become the hot topic of various reviews [14][15][16][17], and special issues of many reputed journals [18,19].
Recently, the advanced technologies make platforms for acquiring the "omics" data generated through numerous experimental sources to come-up with the system level broad perspective of organisms. The genomics and proteomics data helped a lot in knowing cell molecules and their interrelations at the molecular level, so one can determine the biological activity of those components [20]. In this direction, the initial approach is to construct the dataset for metalloproteins that can be used with other proteomics data for the modulation of multifaceted cellular processes integrating metals for which "omics" data could be extremely useful. In the present review, we discuss the recent scientifi c achievements on metallomics and related fi elds advocating the uptake, role, upkeep, and movement of metal ions crucial for living beings. Compared to other omics such as genomics and proteomics, metallomics is a relatively new fi eld; however, they have constructed a tremendous data that can be used to study metallomics. Metallomics must be the core focus of scientifi c investigation because functional genomics and proteomics cannot be completed without the role of metalloenzymes and metal ions [21].
In order to understand the science of ''metallomics'', it is essential to use a novel way to look at the relevance of metals in organisms and as the fraction of ecosystem thus bringing together the environment and cellular life (Singh et al. 2011).
The molecular basis for many metal-dependent biochemical processes even now remains ambiguous [3]. Mechanism of metal sensing, metal uptake and storing or incorporation of metals as a cofactor in the cell is few of the thrust areas to work or to meet the coming challenges in metallomics.
Hence, it is crucial to interpret the signifi cance of metals in governing various biological processes and stress responses by systematic evaluation of metal ions, speciation, and localization in various tissues or under particular conditions. In this report, ''metallomics'' is suggested as a novel scientifi c area emphasizing the importance of research in biometals.

Metals in an environment and their biological significance
Metals in the environment are generally benefi cial but can become a source of toxicity if they exceed a certain limit [22]. With increasing pollution, metal toxicity in the water and soil is constantly growing. The environmental pollutants can be the inorganic, organic or heterogeneous combination of both. Inorganic pollutants from anthropogenic activities include carcinogenic heavy metals for instance arsenic (As), mercury (Hg), nickel (Ni) and many others [23]. The sources of environmental pollution include extermination and improvisation activities, mineral processing and mining, agriculture-related activities, wind-blown dust, sea spray, transportation activities and automobiles and other anthropogenic operations [24]. The environmental contamination by inhabitance of deleterious metals pose threat to living beings and ecological community through direct consumption or contact with polluted soil, the food web, imbibition of infected groundwater, attrition in food quality, decrement in land adoption for agriculture thus eliciting food insecurity, and land occupancy disagreements [25]. Although metals are deleterious when present in excess amount but are also essential in order to perform important processes of life [22]. Since the harmful consequences of metals are widely known, the present review emphasizes the signifi cant and crucial aspects of metals in the living system.

Evolution of metal biology from prokaryotes to eukaryotes
Millions of years ago, environmental conditions were reductive and therefore drastically different from today [26]. The primitive cells being anaerobic could not utilize all the elements accessible due to the presence of a substantial volume of hydrogen sulphide (H 2 S) in the environment [27].
The archean ocean which was reductive and anoxic was rich in iron (Fe), manganese (Mn), and cobalt (Co), thus far low in copper (Cu), zinc (Zn) and molybdenum (Mo) [28]. Therefore, the earliest organisms utilized metals that were abundant in the earliest ocean, Mn, Fe, and Co. The composition of the primitive ocean determined the metal-binding architectures evolution and the organisms' choice of elements [29]. However, as organisms evolved the presence of metals determined the metals inside the cell which further became available to perform the biological functions. Anaerobes utilize part of Co(B 12 ) and Ni as reducing catalysts and Fe as the electron transferring agent while Mg ions are used in catalysis of weak acids. Initially, aerobic bacteria developed utilizing Mo and Cu due to sulfi des oxidation. The cell utilized copper ions in the periplasmic space where they operated on novel substrates like Nitric Oxide produced by oxygen [30]. The prokaryotic cells largely utilized Fe and Mg ions in various metabolic pathways and assisted transcription factors as messengers. However, there was no involvement of metal ions such as Ca, Zn and Cu [31]. Reductive environment supported the evolution of prokaryotic organisms when certain transition metals became available. During the advancement of evolution, this condition, additionally with the redox chemical activity of transition metals culminated in the small free cytosolic compartments in prokaryotes [32]. In order to maintain the moderate metal ion accumulation in the cytosol and to defend the cell against cytotoxic augmentation effects of these intracellular metal ions, modern prokaryotic cells evolved new metal binding proteins [33]. This state of proportionately low metal ions in the cytosol is observed in the present day eukaryotes. However, increased abundance and availability of metal ions due to increased oxygen availability was conducive for eukaryotic organism's evolution [34]. For messenger systems, catalysts, DOI: http://dx.doi.org/10.17352/ojps.000008 structure etc. both metals and non-metals are crucial but their introduction into cellular activity at the comparable amount at the same period of evolution is an uncommon phenomenon. The changing environment leads to this evolution [32].
The change was slow as Oxygen was removed at fi rst by the ferrous and sulfi de in the sea. The need was for organisms to acclimatize to the new environment and they were able to adapt due to the multi-chambered bigger, gradual reproducing, cell system, and the eukaryotes, simultaneously with fastreproducing prokaryotes [35]. The main applicability of additional compartments is the isolation of reductive reactions from the oxidative reactions of the cytoplasm and ions storage vesicles of the cytoplasm was not allowed. The bigger cells possessed fi laments, had an extensible membrane and integrated aerobic bacteria and other symbiotic chambers. An elementary and benefi cial modulation of messenger inter-communication was necessary among the non-fl exible compartments to provide homeostasis and from the environment to provide an elevated sensitivity to cellular surroundings [32]. The Ca ions were utilized in essential messengers, Zn ions in transcription agents and Cu ions generally in vesicles [36]. The Zn and Cu became more accessible as their sulphides can be partially oxidized.
Anaerobic cells acquired defensive mechanisms so as to survive in the oxidizing environment. In addition, cell mastered to use oxygen for respiratory metabolism, thereby augmenting the effi cacy of ATP synthesis [37]. For instance, the role of superoxide dismutase and catalase is well demonstrated in protecting the cell against the toxic residues of oxygen metabolism. Cells made use of at least some metals that today are called the "essential" metals. Due to the adequacy of essential metals at the time of prokaryotic and subsequently eukaryotic organism's evolution many metals integrated with biochemical functions as elementary factors [38] (Figure 1).

Emergence of novel omics: Metallomics
The fi elds like genomics, proteomics, and metabolomics have collectively known as "omics" had been subjected to tremendous improvement in the past. The fi rst to develop was genomics, which produces complete genomic DNA sequences of living creatures. The next "omics" includes proteomics i.e. the interpretation of the structure, stability, localization, and interaction of cellular or organism's proteins [39].
Further, metabolomics annotates the complete metabolites of an organism [40]. The techniques used in proteomics and metabolomics have usually evaded the metals present in proteins and metabolites. However, the cell chemistry should be characterized not only by its nucleic acids, proteins, and metabolites but also with its metallome [16]. Therefore, metallomics should be recognized as emerging omics that characterize the metallomes and fi nd out the interactions and functional associations of metals with proteins, genes, and metabolites [11]. The high-throughput technologies provided tremendous data about the genome. "Omics" data are accessible for many cell constituents and interactions, in addition to, for instance, genomics, proteomics, transcriptomics and proteinprotein interaction maps (interactomics) [41]. The availability of large-scale information has largely helped researchers in interpreting functions of organisms but due to inadequate metallome data, the understanding remains partial. However, to explore any biological function/signaling pathway/cellular mechanism, metallomics should be linked with other omics and so in this review further we explain how other omics require metallomics to retrieve any information.

Integration of metallomics with genomics
The tremendous advancement in omics technology has led to the growth of genomics to such an extent that, it is divided into structural genomics and functional genomics. Structural genomics procreates high-resolution, 3D structural models of DNA whereas functional genomics tries to decipher functions of various genes through RNA analysis [42]. The metallomics and metalloproteomics are comparatively novel areas of study; however, data generated by genomics can be utilized to rapidly enhance our perceptive of metalloids and metalloproteomics [43].   [46]. The lack of cations in several of such motifs leads to damaged or more fl exible tertiary structure [47]. Additionally, to stabilize the junctions in DNA such as transitional Holliday junction in genetic recombination, the divalent ions mainly magnesium is crucial. The magnesium ions in the junction shield the negatively charged phosphate groups. This helps to position the phosphate groups adjacent to one another conceding stacked conformation [48].
The several metalloenzymes and metal ions assist in the synthesis and the metabolic roles of genes and proteins.
The amplest transition metal, Zn, aids in carrying out the activity of over three hundred enzymes, stabilization of DNA and expression of genes [49]. The zinc in the eggs of salmon probably assists in the synthesis of DNA/RNA, processes such as the production of energy adenosine triphosphatase (ATPase) and regulation of cell division. Further, calcium ions associate with the carbonyl terminals in the proteins so as to stabilize the structure. Therefore, the imaging of metallomics will concede co-localization of gene expression as well as localization of protein patterns with metals thus providing gene, protein, and metal function linkage [8]. Unlike the correlation of metallo-proteome and genome, the correlation of metallometabolome and genome is not so distinct and may be missing.
With the molecular cloning methods the metallo-metabolome interaction can be established by investigating metal resistance genes in organisms [11]. Therefore, the three -omics are correlative, and the aggregation of authenticated metallomic data with the transcriptome and proteome knowledge of a cell is the largest challenges for future investigation in this area. One way forward is that better established -omics (transcriptomics and proteomics) incorporate metallomics in their studies on model organisms whose gene/protein sequences are well annotated in databases.

Metallomics interaction with proteomics
The metalloproteins three-dimensional structure incorporates inorganic ions and comprises nearly 30% of the proteins [50]. Metalloproteins catalyze signifi cant processes of water oxidation and photosynthesis [51]. These are mostly two-electron redox mechanisms and involve atom or group transfers. The oxygen atom is added to the substrate in many important and common reactions.  [66].
Metallochaperones guide the ions and defend it from the over-chelation potentiality of the cell [67]. Metallochaperones sort specifi c metal co-factor to the precise metalloenzymes and transport them to diverse location. Many metal ions present in the enzymes acting as co-factors remain located intracellularly or transported extracellularly [68]. For instance, copper is toxic element yet crucial for the living systems. Cells acquire many mechanisms so as to sustain copper homeostasis such as; the protein-mediated intracellular delivery of copper to target proteins. This is consummated by a group of proteins, the copper chaperons, which conserved proteins present in prokaryotes and eukaryotes. Three different classes of Cumetallochaperones namely cytochrome oxidase (COX17), antioxidant (ATX1/ HAH1) and copper chaperones of SOD1 (CCS) are known [69]. The copper ion incorporation into SOD1 requires a Cu metallochaperone [67]. Cd (II), Zn(II) and Cu(I) in metal clusters [73]. Through the thiols present in cysteine residues, MTs bind the physiological (Cu, Zn Se) and xenobiotic (Ag, As, Cd, Hg) heavy metals [74].
The metal clusters Me(II) 3 (Cys) 9 and Me(II) 4 (Cys) 11 are present in mammalian MTs. These clusters bind heptad of metal ions (Me) through thiolate coordination [75]. Biosynthesis of MTs is induced by several agents and is coordinated at the level of transcription. MTs sequester mainly metals which are nonessential [76].

Metallomics role in metabolomics
Metabolomics, a rapidly developing technology deals with metabolome which is described as the metabolites compilation of the cell [77]. Metabolic fi ngerprinting, targeted analysis and metabolite profi ling are the major approaches presently DOI: http://dx.doi.org/10.17352/ojps.000008 adopted in metabolomics related research [78]. The three components are signifi cant in metabolomics in order to carry out such processes: (1) antioxidants (2) stress by-products due to disruption of homeostasis; and (3) molecule involved in signal transduction responsible for adaptation response. The molecules involved in signal transduction are either recurrently synthesized or compounds liberated from conjugated forms such as salicylic acid [79]. The metals and ligands interaction is a signifi cant area of metallomics research. The defensive mechanisms of numerous organisms against toxic metals comprise of many organic acids such as succinate, malate, oxalate etc. [16], metallophores [80], and peptides binding metals [81]. Hyper accumulating plants developing metal homeostasis are of particular interest. They can live and reproduce in metal-rich environments [82]. In such plant cells, the complexity of metals leads to many relatively poorly characterized metal complexes.
The study of detoxifi cation controlling mechanisms can benefi t from the establishment of the species formed. The metabolite and gene networks can be deciphered through assimilation of the data of metabolomics and transcriptomics [83]. For example, the proteomics and the metabolomics were integrated to establish the response of Arabidopsis to cesium stress [84]. However, in view of the problem of data integration, the collective study of the data from metabolomics and other omics is actually challenging [85]. Though other targets for approaches related to genomics are attainable, to identify metabolites produced in streptomycetes grown in growth media with 0.2 mM Ni or Cd indicates huge probability for revealing strains from cultures. This would help to decipher cluster of genes during metal stress [86]. Metals could be used as inducers of new metabolites such as antibiotics i.e. another area of metallo-metabolomics used in biotechnology [14].
A good approach to metallo-metabolomics should help in element identifi cation in the species or identifi es at least some and account clearly for the non-identifi ed ones (quantitative speciation blueprint). In contrast to metallo-proteomics, the de novo recognition of metabolites is performed by using leading spectrometric techniques. Mass spectrometry (MS) technology and a purifi cation method are a comprehensive approach to metallo-metabolomics [11]. Metallomics comprise various types of information from identifi cation of metals (qualitative metallomics) to fi nding out their level (quantitative metallomics) [14]. Metallomics may be important in studies relevant to plant metal tolerance and homeostasis [93], biogeochemical metal cycles [94], plant proteome annotation engaged in toxicity of metals [95]. Metallomics may be supportive to the improvement of applications for optimized strategies in metal contaminated soils, including microbial-assisted phytoremediation of polluted land, reclamation of soil and biomarker recognition for eco-toxicological studies [94].
For phytoremediation studies, phytochelatins build-up in the plants is signifi cant phenomenon due to their role in detoxifi cation of heavy metals [96]. The study of plant and soil amount of heavy metals in Opuntia fi cus predicated a complex interplay of heavy metals in the environment and phytochelatin production [97]. Many advanced approaches such as HPLC-

ICP-MS is utilized in phytoremediation studies for the establishment of seleno-compounds in the plant extracts from
Brassica juncea, Allium sativum, [98], and Bertholletia excelsa [99].
In in the metabolism of the cell and assist in identifi cation of metal cofactor present in the protein. This helps to fi nd out the function of the protein in cellular pathways [100]. Many metalloproteins act as organism's markers, exposure to metals and indicators of their status. For example, metallothioneins are metal binding proteins rich in Cys and are utilized as biomarkers for some organisms exposed to metals in the environment.
Metallothioneins in the organisms bind metals and thus act as a detoxifying mechanism [101]. Metallothioneins are utilized to fi nd out the effect of heavy metals in marine organisms [102]. The

Metallomics in communication biology
Metals not only perform an important function in various crucial processes of life but they are also used as magnetic compasses in living organisms. The various animals such as bees, birds, sea turtles etc. possess compasses inside them. With their compasses, some species navigate entire oceans [105].
Magnetoreception or magnetoception allows the animals which show biomagnetism (magnetic fi eld production by organisms) to investigate magnetic fi eld so as to perceive directions.
Magnetoception illustrates the navigational skills of many animal species [106].

Metallomics in pharmacy / medicine
Metal complexes are important to carry out biological and biomedical processes. Metal complexes consist of meal atoms surrounded by ligands. Organic compounds in medicine have different modes of action; some are biotransformed by metalloenzymes (Holm and Solomon 1996) others have an effect on the metabolism of metals. Metal complexes are used as drugs for curing various human diseases like lymphomas, carcinomas, infection control, anti-infl ammatory, diabetes, and olfactory disorder. The transition metals upon interaction with molecules having negative charges display different oxidation states. This phenomenon of transition metals is signifi cant for the advancement of drugs based on metals having therapeutic values and pharmacological applications [111]. The metal complexes signifi cance in pharmacy and the recent developments are indexed in table 2.
Novel therapeutics with metal complexes offers real possibilities to pharmaceutical industries. The smooth encounter of the metal complexes in ligand substitution and redox reactions presumably indicate active species to be the biotransformation products of the administered complex [112].
Therefore, metal compounds could be used more effectively as drugs following the active species identifi cation. The metal complexes identifi cation and their biotransformation still require investigation. Further, the toxicity and metal compounds are correlated therefore targeting metal-based drugs to specifi c locations (cells, tissues or receptors) where they are needed, could lower the toxicity to a signifi cant extent [113]. Thus, to understand bio-coordination chemistries of drugs having metal ions, thermodynamics and kinetics of metal complex reactions, mainly under biologically relevant conditions is signifi cant [114]. Hence, the study of elemental medicine would help to decipher diagnostic and therapeutic approaches and contribute to our insight of natural biological processes.

Contribution of bioinformatics in exploration of metallomics
The huge repository of genome sequence data is responsible for the boom in bioinformatics. Although the information of metalloproteome is required for the comprehensive understanding of life processes, the presently available DOI: http://dx.doi.org/10.17352/ojps.000008 experimental methods or techniques are not capable of achieving such a demanding task. At this point, bioinformatics provides a considerable contribution to subjugate the limitation of empirical methods by using predictive tools and information technology [115]. The metal binding domains can be deciphered with the help of protein sequence data available in Pfam and other libraries [116]. Bertini and Cavallaro [117], have reviewed the bioinformatics approaches dedicated to bimetals. Many metalloprotein databases using primary sequence information have been generated utilizing various bioinformatics methods [118].
With the advancement in high throughput techniques the characterization of metallome would be easy and thus the huge amount of data will be obtained. Therefore, bioinformatics approaches would be required to retrieve the data from the tremendous data set. The computational program utilizes sequence information by taking into count position-specifi c evolutionary profi les and aspects such as protein length and amino acid composition. This method provides information on transition metal binding site components and is highly complementary to the high-throughput techniques based on X-ray absorption spectroscopy (HT-XAS), which identifi es the metal binding to protein [119]. Further, comparative structure modeling is a very powerful tool in driving protein function.
MODBASE is structural models data [120] and is now discontinued [122]. Other related databases include

Conclusion and Future Prospective
Metallomics is projected as the interdisciplinary research