322,944 research outputs found
Soil health and arthropods: From complex system to worthwhile investigation
No full textThe dramatic increase in soil degradation in the last few decades has led to the need to identify methods to define not only soil quality but also, in a holistic approach, soil health. In the past twenty years, indices based on living communities have been proposed alongside the already proven physical-chemical methods. Among them, some soil invertebrates have been included in monitoring programs as bioindicators of soil quality. Being an important portion of soil fauna, soil arthropods are involved in many soil processes such as organic matter decomposition and translocation, nutrient cycling, microflora activity regulation and bioturbation. Many studies have reported the use of soil arthropods to define soil quality; among taxa, some have been explored more in depth, typically Acari and Collembola, while generally less abundant groups, such as Palpigradi or Embioptera, have not been investigated much. This paper aims to evaluate and compare the use of di_erent soil microarthropod taxa in soil degradation/quality studies to highlight which groups are the most reported for soil monitoring and which are the most sensitive to soil degradation. We have decided not to include the two most present and abundant taxa, Acari and Collembola, in this paper in consideration of the vast amount of existing literature and focus the discussion on the other microarthropod groups. We reported some studies for each taxon highlighting the use of the group as soil quality indicator. A brief section reporting some indices based on soil microarthropods is proposed at the end of this specific discussion. This paper can be considered as a reference point in the use of soil arthropods to estimate soil quality and health
Understanding the mechanism of ZinT-mediated metal acquisition: a thermodynamic study
No full textZinT is a periplasmic protein found in Gram-negative bacteria. It is involved in cellular metal trafficking and may function as zinc-chaperone for the ZnuABC transporter. There is a general consensus that the three highly conserved histidine residues (His167, His176 and His178) facing the centre of ZinT calycin like-domain and the amino-terminal fragment between residues 24 and 29 (-HXHHXH-) should be effective zinc binding sites [1, 2]. The Zn2+-ZinT complex from S. enterica can interact with ZnuA forming a ternary complex where both proteins expose their binding pocket to the Zn2+ ion and where the His-rich loop of ZnuA functions as a hypothetical metal transfer intermediary between the two proteins [3, 4]. The main aim of this work is therefore to provide insight into the correlation between the metal-binding ability of ZinT and its biological role. The chosen unstructured fragments, which serve as models to simulate the coordination and transport of metal ions in ZinT protein (see Figure 1) from Escherichia coli and Salmonella enterica, correspond to the 24–29 and 166–178 amino acid sequences and are protected at the amino- and carboxyl-termini: Ac-24HGHHSH29-Am and Ac-166DHIIAPRKSSHFH178-Am (E. coli), Ac-24HGHHAH29-Am and Ac-166DHIIAPRKSAHFH178-Am (S. enterica). Interestingly, both the metal-binding sites of ZinT from S. enterica undergo a Ser-to-Ala substitution (position 28 and 175). A deep investigation of the complex-formation equilibria and coordination chemistry of the formed species has been performed through different experimental techniques, including potentiometry, mass spectrometry and various spectroscopies.
The obtained results highlight novel insight into the mechanism of ZinT-mediated metal acquisition and allow a comparison with other biologically relevant metal-binding systems, such as the antimicrobial peptide calcitermin which can, in principle, participate in human nutritional immunity, competing with ZinT for the metal ion acquisition.
Financial support of the National Science Centre (UMO-2017/26/A/ST5/00364 and UMO-2020/37/N/ST4/03165) is gratefully acknowledged. This paper is based upon work from COST Action CA18202, NECTAR – Network for Equilibria and Chemical Thermodynamics Advanced Research, supported by COST (European Cooperation in Science and Technology).
References:
[1] J. Chen, L. Wang, F. Shang, Y. Dong, N.-C. Ha, K. H. Nam, C. Quan, Y. Xu, Biochem. Biophys. Res. Commun. 2018, 500(2), 139-144.
[2] H. G. Colaço, P. E. Santo, P. M. Matias, T. M. Bandeiras, J. B. Vicente, Metallomics 2016, 8(3), 327-336.
[3] A. Ilari, F. Alaleona, G. Tria, P. Petrarca, A. Battistoni, C. Zamparelli, D. Verzili, M. Falconi, E. Chiancone, Biochim. Biophys. Acta 2014, 1840(1), 535-544.
[4] D. Bellotti, M. Rowińska-Żyrek, M. Remelli, Dalton Trans. 2020, 49(27), 9393-9403
The impact of metal coordination on calcitermin antimicrobial properties
No full textCalcitermin (VAIALKAAHYHTHKE) is a 15-mer antimicrobial peptide found in human nasal fluid [1] which is of particular interest thanks to its metal chelating ability. Noteworthy, it exhibits improved antifungal (C. albicans) and antibacterial properties in the presence of Zn2+ and Cu2+ ions under acidic conditions, while the histidine-to-alanine mutation in position 9, 11 and 13 can modulate the activity against certain microorganisms [2].
In light of the above, we decided to extend the study to other calcitermin derivatives, where the amino acid sequence is modified to understand the impact of metal coordination on the antimicrobial activity. The new synthetised derivatives include the C- and/or N- terminal protected peptides, which could also decrease the susceptibility towards exopeptidases, the alanine-to-serine mutants (A7S, A8S and A7/8S), which are designed to stabilize copper complexes [3] and the truncated analogue Ac-AHYHTHKE-NH2 to verify if the metal coordination site of natural calcitermin can correspond to the minimum active sequence.
This work is therefore aimed at characterizing the interaction of Zn2+ and Cu2+ ions with calcitermin derivatives in aqueous solutions. A deep investigation of the thermodynamic parameters of complex formation equilibria and of the coordination chemistry of the formed species has been obtained by means of several techniques, including potentiometry, high-resolution mass spectrometry, UV-Vis, circular dichroism and EPR.
Financial support of the Polish National Science Centre (UMO-2020/37/N/ST4/03165) and of the COST Action CA18202, NECTAR – Network for Equilibria and Chemical Thermodynamics Advanced Research is gratefully acknowledged.
[1] M. Cole, Y.-H. Kim, S. Tahk, T. Hong, P. Weis, A.J. Waring, T. Ganz, T. FEBS Lett 504 (2001) 5-10.
[2] D. Bellotti, M. Toniolo, D. Dudek, A. Mikołajczyk, R. Guerrini, A. Matera-Witkiewicz, M. Remelli, M. Rowińska-Żyrek, Dalton Trans. 48 (2019) 13740-13752.
[3] D. Bellotti, A. Miller, M. Rowińska-Żyrek, M. Remelli, Biomolecules 12 (2022) 121
Study of Cu(II) and Zn(II) interaction with the metal binding domain of ZinT protein
No full textUnderstanding the mechanism of metal trafficking at the host/pathogen interface can help designing innovative antibiotic therapies. In fact, bacteria rely on sophisticated systems (e.g. metallophores) to sequester metals from the host environment during infections. In the attempt to investigate these metal-acquisition processes, we studied the complexation of Cu(II) and Zn(II) – two endogenous and competing metal ions – with some peptide fragments of ZinT, a periplasmic protein found in several bacterial species and mostly involved in Zn(II) recruitment. Its most probable metal-binding site corresponds to a domain containing three histidine residues (positions 167, 176 and 178) and one aspartic acid (position 166, Fig. 1). ZinT also possesses a highly conserved histidine-rich loop (HGHHXH, residues 124-129), whose participation in metals uptake has also been suggested [1,2].
By means of ESI-MS, potentiometry, UV-Vis/CD spectrophotometry and EPR measurements, we studied the formed metal complexes with the protected peptides Ac-124HGHHSH129-Am and Ac-166DHIIAPRKSSHFH178-Am (of ZinT sequence from Escherichia coli), and Ac-124HGHHAH129-Am and Ac-166DHIIAPRKSAHFH178-Am (ZinT-Salmonella enterica). We ultimately compared ZinT with some human-defence mediators, e.g. the antimicrobial peptide Calcitermin [3], to evaluate the metal effectiveness in the expression of the pathogenic/antimicrobial activity by the studied systems.
[1] A. Ilari, F. Alaleona, G. Tria, P. Petrarca, A. Battistoni, and C. Zamparelli, Biochim. Biophys. Acta. 1840(1) (2013) 535–544.
[2] P. Petrarca, S. Ammendola, P. Pasquali, and A. Battistoni, J. Bacteriol. 192(6) (2010) 1553–1564.
[3] D. Bellotti, M. Toniolo, D. Dudek, A. Mikolajczyk, R. Guerrini, A. Matera-Witkiewicz, M. Remelli, and M. Rowinska-Zyrek, Dalton Trans. 48(36), (2109) 13740–13752
Understanding the thermodynamics and coordination chemistry of metal-binding proteins: the common thread to elucidate metal acquisition processes at host/pathogen interface
No full textSeveral studies have shown that the disruption of metal homeostasis in bacterial and fungal cells can be a powerful tool to design new antimicrobial drugs with a high rate of selectivity and specificity. To prevent infections, in fact, the human organism reduces the bioavailability of essential micronutrients by means of an innate immune response termed “nutritional immunity”; on the contrary, pathogens rely on specialized metal-binding proteins and molecular systems which capture the metal ions from the competitive host environment forming stable complexes [1].
Understanding the properties, structure and action mechanisms of the involved metal chelators is the very first step to elucidate the dynamics behind the metal transfer mechanisms and to rationally design novel metal-based antibiotic therapeutics [2].
An outstanding example is given by the thermodynamic and spectroscopic characterization of the zinc and copper binding sites of the periplasmic protein ZinT, expressed by Escherichia coli and Salmonella enterica [3, 4]. The chosen unstructured fragments, which serve as models to simulate the coordination and transport of metal ions in ZinT protein, correspond to the 24–29 and 166–178 amino acid sequences and are protected at their amino- and carboxyl-termini: Ac-24HGHHSH29-Am
and Ac-166DHIIAPRKSSHFH178-Am (E. coli), Ac-24HGHHAH29-Am and Ac-
166DHIIAPRKSAHFH178-Am (S. enterica).
A deep investigation on the thermodynamics and coordination chemistry of the formed Zn(II) and Cu(II) complexes was performed through different experimental techniques. The protonation and complex-formation equilibria were studied by means of potentiometric acid-base titrations. ESI mass spectra of the solutions under examination allowed to confirm the stoichiometries of the formed species and, through UV-Vis, CD and EPR spectroscopies at variable pH values, the metal
coordination spheres and the geometry of the complexes were explored. Finally, the obtained results allowed a comparison with other biologically relevant metal-binding systems, such as the antimicrobial peptide calcitermin (VAIALKAAHYHTHKE) which can, in principle, participate in human nutritional immunity, competing with ZinT for the metal ion acquisition.
Financial support of the National Science Centre (UMO-2020/37/N/ST4/03165) is gratefully acknowledged. This paper is based upon work from COST Action CA18202, NECTAR – Network for Equilibria and Chemical Thermodynamics Advanced Research, supported by COST (European Cooperation in Science and Technology).
References
[1] D. A. Capdevila, K. A. Edmonds, D. P. Giedroc, Essays Biochem., 2017, 61(2), 177-200.
[2] S. R. Hennigar, J. P. McClung, Am. J. Lifestyle Med., 2016, 10(3), 170-173.
[3] A. Battistoni, A. Ammendola, E. Chiancone, A. Ilari, Future Med. Chem., 2017, 9(9), 899-910.
[4] D. Bellotti, M. Rowińska-Żyrek, M. Remelli, Dalton Trans., 2020, 49(27), 9393-9403
Unrevealing the antimicrobial properties of calcitermin and its peptide derivatives
No full textThanks to the broad spectrum of activity and scarce attitude to induce antimicrobial resistance, antimicrobial peptides (AMPs) represent a rational chance to overcome the current drug-resistance crisis. Among several uncharacterized molecules that contribute to the overall antimicrobial activity of human nasal fluid, a 15-residue antimicrobial peptide named calcitermin (VAIALKAAHYHTHKE) has been identified [1].
Calcitermin contains a metal-binding domain with three alternated histidine residues (His9, His11 and His13) and the free terminal amino and carboxyl groups. Based on our preliminary studies, it exhibits an improved microbicidal activity when Zn2+ or Cu2+ ions are present in the culture medium. Additionally, calcitermin His-to-Ala mutants – where each histidine residue is replaced with one alanine – have different metal coordination modes, resulting in significant changes of the antimicrobial properties [2].
These promising results prompted us to focus on calcitermin derivatives where the peptide structure is modified in order to confer higher proteolytic stability. The first task of this work consists of a careful evaluation of the enzymatic stability of native calcitermin in human plasma. Afterwards, C- and/or N- terminal modifications have been introduced to possibly obtain calcitermin derivatives resistant to proteases [3]. Changes in the peptide backbone, and in particular the N-terminus protection, can affect the calcitermin metal-binding behaviour and therefore further investigations on the metal interaction with the synthesized protected peptides have been performed, in order to connect the antimicrobial activity of calcitermin with the complex-formation ability. The characterization of metal complexes has been performed by means of several techniques, including potentiometry, high-resolution mass spectrometry, NMR, UV-Vis, circular dichroism and EPR. The obtained results will allow us to propose and design new therapeutic antimicrobial strategies based on calcitermin derivatives and their metal complexes.
Financial support of the Polish National Science Centre (UMO-2020/37/N/ST4/03165) and of the COST Action CA18202, NECTAR – Network for Equilibria and Chemical Thermodynamics Advanced Research is gratefully acknowledged.
[1] M. Cole, Y.-H. Kim, S. Tahk, T. Hong, P. Weis, A. J. Waring, and T. Ganz, FEBS Lett. 504 (2001) 5-10.
[2] D. Bellotti, M. Toniolo, D. Dudek, A. Mikołajczyk, R. Guerrini, A. Matera-Witkiewicz, M. Remelli, and M. Rowińska-Żyrek, Dalton Trans. 48 (2019) 13740-52.
[3] X. Lai, J. Tang, and M. E. H. ElSayed, Expert Opin. Drug Discov. (2021) 1-16
Zincophore metal binding sites: from solution equilibria to metal transport in human pathogens
No full textThe assimilation of metal nutrients from the host environment is an important factor for the onset and progression of infectious diseases. Zn(II) ions, in particular, are crucial for the virulence and survival of both pathogen and human cells, being indispensable for the expression and function of many enzymes. Interestingly, the Zn(II) uptake mechanism is one of the major differences in the metabolism of human and pathogen cells, thus it represents a promising drug-target for specific and selective treatments [1].
Pathogens compete with the host organism in order to acquire zinc and meet their physiological metal nutrient demands. An efficient zinc recruitment is therefore achieved by means of specialized zinc-binding proteins and molecular systems which capture the metal ion from the host environment forming stable complexes [2, 3]. A deep knowledge of the properties, structure and action mechanisms of these extracytoplasmic zinc chelators (zincophores) can be a powerful tool to design new therapeutic strategies against the antibiotic and/or antifungal resistance [4].
As a first step, it is essential to obtain information about thermodynamics and coordination chemistry of the involved systems, in order to point out the most effective metal binding sites and to elucidate the dynamics behind the metal transfer. Indeed, a relatively high metal binding affinity is crucial to ensure the acquisition process in an environment rich of competitive systems and thermodynamic studies may help clarifying this aspect. An outstanding example is given by the characterization of the zinc-binding sites of the periplasmic protein ZinT, expressed by Escherichia coli and Salmonella enterica [5].
References:
[1] Capdevila, D.A.; Edmonds, K.A.; Giedroc, D.P., Metallochaperones and metalloregulation in bacteria. Essays Biochem., 2017, 61 (2), 177-200.
[2] Hennigar, S.R.; McClung, J.P., Nutritional Immunity: Starving Pathogens of Trace Minerals. Am. J. Lifestyle Med., 2016, 10 (3), 170-173.
[3] Morey, J.R.; Kehl-Fie, T.E., Bioinformatic Mapping of Opine-Like Zincophore Biosynthesis in Bacteria. mSystems, 2020, 5 (4), e00554-00520.
[4] Battistoni, A.; Ammendola, S.; Chiancone, E.; Ilari, A., A novel antimicrobial approach based on the inhibition of zinc uptake in Salmonella enterica. Future Med. Chem., 2017, 9 (9), 899-910.
[5] Bellotti, D.; Rowińska-Żyrek, M.; Remelli, M., Novel insights into the metal binding ability of ZinT periplasmic protein from Escherichia coli and Salmonella enterica. Dalton Trans., 2020, 49 (27), 9393-9403
Calcitermin and its peptide derivatives as promising antimicrobial agents with metal chelating ability
No full textThanks to the broad spectrum of activity and scarce attitude to induce antimicrobial resistance, antimicrobial peptides (AMPs) represent a rational chance to overcome the current drug-resistance crisis. Among several uncharacterized molecules that contribute to the overall antimicrobial activity of human nasal fluid, a 15-residue antimicrobial peptide named calcitermin (VAIALKAAHYHTHKE) has been identified [1]. Calcitermin contains a metal-binding domain with three alternated histidine residues (His9, His11 and His13) and the free terminal amino and carboxyl groups (Figure 1). It also exhibits an improved microbicidal activity when Zn2+ or Cu2+ ions are present in the culture medium [2]. Additionally, calcitermin His-to-Ala mutants – where each histidine residue is replaced with one alanine – have different metal coordination modes, resulting in significant changes of the antimicrobial properties. These results prompted us to focus on calcitermin derivatives where the peptide structure is modified to confer higher proteolytic stability but maintaining the metal chelating ability. Therefore, C- and/or N- terminal modifications have been introduced to possibly obtain calcitermin derivatives resistant to proteases [3]. Changes in the peptide backbone can affect the metal-binding behaviour and therefore further investigations on Zn2+ and Cu2+ interaction with calcitermin analougues are required to connect the antimicrobial activity with the complex-formation ability. The characterization of metal complexes has been performed by means of several techniques, including potentiometry, high-resolution mass spectrometry, UV-Vis, circular dichroism, NMR and EPR. The obtained results will allow us to propose and design new therapeutic antimicrobial strategies based on calcitermin derivatives and their metal complexes. Financial support of the Polish National Science Centre (UMO-2020/37/N/ST4/03165) is gratefully acknowledged.
References
[1] Cole, A. M., Kim, Y.-H., Tahk, S., Hong, T., Weis, P., Waring, A. J., Ganz, T. Calcitermin, a novel antimicrobial peptide isolated from human airway secretions. FEBS Lett., 504 (2001) 5-10.
[2] Bellotti, D., Toniolo, M., Dudek, D., Mikołajczyk, A., Guerrini, R., Matera-Witkiewicz, A., Remelli M., Rowińska-Żyrek, M., Dalton Trans., 48 (2019) 13740-13752.
[3] Vlieghe, P., Lisowski V., Martinez J., Khrestchatisky M. Synthetic therapeutic peptides: science and market. Drug Discov. Today, 15 (2010) 40-56
Macrochelates from rigid polypeptides: a step forward towards synthetic enzymes?
No full textsimple method to obtain branched peptides consists of binding a well-defined number of
peptide sequences to a central scaffold. Molecules of this type have been synthesized in the
past and have been investigated as new anticancer drugs, vaccines and antibacterial agents
[1]. In this way, if the peptides used to build up these structures have binding properties
towards metal ions, it is possible to obtain completely new polydentate peptide ligands, with
the ability to mimic the coordination sites of natural metalloenzymes.
In this context, the present contribution reports the results of a pilot study in which:
first of all, the two oligopeptides AAHAWG-NH2 (P1) and HAWG-NH2 (P2), protected at their
carboxylic terminus, were synthesized and studied as ligands of Cu(II) and Zn(II); secondly,
the corresponding tetrameric branched forms were synthesized and investigated. The latter
were obtained by binding four identical peptides, of one of the two kinds, to a cyclam
platform, through a cysteine-maleimide bridge (Scheme 1) [2].
While the binding behaviour of P1 and P2 towards the considered metal ions is
perfectly in line with what already known in the literature for oligopeptides that contain a
histidine residue in position 3 (ATCUN-type) or 1 (histamine-like) [3,4], in the case of
tetrameric structures, peculiar behaviours are observed. In fact, the branched peptides
contain four imidazole residues which are blocked, through a spacer, to a single cyclam
platform, resulting very close each other and thus producing a very high local concentration
of the donor atoms.
References:
[1] Pini, A.; Falciani, C.; Bracci, L., Branched Peptides as Therapeutics. Curr. Protein
Pept.Sci.2008, 9, 468-477.
[2] Guerrini, R.; Marzola, E.; Trapella, C.; Pelà, M.; Molinari, S.; Cerlesi, M. C.; Malfacini, D.;
Rizzi, A.; Salvadori, S.; Calò, G.; A novel and facile synthesis of tetra branched
derivatives of nociceptin/orphanin FQ. Bioorg. Med. Chem.2014, 22, 3703–3712.
[3] Zamariola G.; Watly J.; Gallerani E.; Gavioli R.; Guerrini R.; Kozlowski H.; Remelli M.;
AGHLDDLPGALSAL: a hemoglobin fragment potentially competing with albumin to bind
transition metal ions. J. Inorg. Biochem.2016, 163, 301-310.
[4] Remelli, M.; Conato, C.; Agarossi, A.; Pulidori, F.; Młynarz, P.; Kozłowski, H.; Copper
complexes of dipeptides with l-Lys as C-terminal residue: a thermodynamic and
spectroscopic study. Polyhedron2000, 19, 2409–2419
Metallo peptides in the design of new, highly selective clinical treatments
No full textThe necessity of new antimicrobial agents is unarguable, since current therapeutic treatments are not
always effective: pathogenic microorganisms capable to adapt and resist against drug action have
significantly increased over the last decades and their associated mortality rate still remains a global
concern. Several studies have shown that metal acquisition and regulation greatly contribute to
virulence and physiology of pathogenic species. In particular, to prevent infections humans restrict
the access to essential micronutrients by means of an innate immune response termed "nutritional
immunity”, on the contrary pathogens rely on sophisticated systems (e.g. siderophores) to overcome
the scarce metal bioavailability. From this perspective, a deeper insight into the mechanism of metal
trafficking in pathogens and host nutritional immune response can provide crucial information to
design new effective antibiotic therapies, e.g. by developing species-selective transport or imaging
drugs, based on metal complexes, which can be recognized only by specialized metal transport
proteins (“Trojan Horse” approach). Furthermore, metal complexes cannot be only a promising tool
in antimicrobial treatments, but they can also find application against pathologies which – in general
– involve metal ions (e.g. neurodegenerative diseases or cancer) [1-3].
As a first step, it is essential to obtain information about thermodynamics and coordination
chemistry of the metal chelators, in order to point out the most effective metal binding sites. Both
natural and artificial peptides can be exploited for this purpose. Aiming at the design of new clinical
treatments, we focused on Zn(II) and Cu(II) binding behavior towards both the putative metal
transporter C4YJH2 (a protein sequence of 199 residues found in the genome of Candida albicans
[4]) and the antimicrobial peptide Calcitermin [5], isolated in the human airways.
The characterization of the complexes required a variety of techniques. The stoichiometry of
the formed species has been determined through high-resolution mass spectrometry as well as by
potentiometry which also provided the stability constants of all the formed metal complexes;
thermodynamic description of metal-ligand interactions has been also investigated by calorimetry.
The identification of binding sites and the coordination geometry of the formed species have been
achieved by several spectroscopic techniques (NMR, UV-Vis, fluorimetry, CD and EPR).
REFERENCES
[1] M. Blatzer, J.P. Latgé, Curr. Opin. Microbiol. 2017, 40, 152-159.
[2] A. Szebesczyk, E. Olshvang, A. Shanzer, P.L. Carver, E. Gumienna-Kontecka, Coord. Chem. Rev. 2016, 327, 84-109.
[3] E. R. Ballou and D. Wilson, Curr. Opin. Microbiol. 2016, 32, 128–134.
[4] D. Bellotti, D. Łoboda, M. Rowińska-Żyrek, M. Remelli, New J. Chem. 2018, 42, 8123-8130.
[5] A.M. Cole, Y.-H. Kim, S. Tahk, T. Hong, P. Weis, A.J. Waring, T. Ganz, FEBS Lett. 2001, 504, 5-10
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