https://pubs.rsc.org/en/content/articlelanding/2007/CC/b709515e#!divAbstract

Inhibition of SARS coronavirus helicase by bismuth complexes†

Nan Yang,^a   Julian A. Tanner,^b   Zai Wang,^b   Jian-Dong Huang,^b   Bo-Jian Zheng,^c   Nianyong Zhu^a  and  Hongzhe Sun*^a  

Author affiliations

Abstract

A series of bismuth complexes were synthesized and characterized, and most of them exhibited inhibition against the SARS coronavirus helicase ATPase and duplex-unwinding activities at micromolar concentrations.

Porphyriineistä etsitään terapeuttisia lääkeaineita monia viruksia vastaan.

Moni porfyriinirakenne on kuitenkin toksinen ja seulontavaiheen tietoja löytyy. 

Yersinia pestis, keuhkorutto indusoituvan HO-1 i:n terapeuttinen potentiaali. Hemianalogi kobolttiprotoporfyriini yriinin(CoPP)

https://aac.asm.org/content/64/4/e01819-19etin tämän myös  ystäville Facebookiin:  keuhkoruton muutoksia vähentämään  ( koehiirillä) on  kehitetty  malli. Hypoteesi:  indusoituvan  hemioksygenaasi1:n  aktivoiminen  hemianalogilla CoPP, kobolttiprotoporfyriinilla,  esti noita keuhkoruttomuutoksia.

Experimental Therapeutics

Activation of Heme Oxygenase Expression by Cobalt Protoporphyrin Treatment Prevents Pneumonic Plague Caused by Inhalation of Yersinia pestis

Joshua L. Willix, Jacob L. Stockton, Rachel M. Olson, Paul E. Anderson, Deborah M. Anderson

DOI: 10.1128/AAC.01819-19

ABSTRACT

Pneumonic plague, caused by the Gram-negative bacteria Yersinia pestis, is an invasive, rapidly progressing disease with poor survival rates. Following inhalation of Y. pestis, bacterial invasion of the lungs and a tissue-damaging inflammatory response allows vascular spread of the infection. Consequently, primary pneumonic plague is a multiorgan disease involving sepsis and necrosis of immune tissues and the liver, as well as bronchopneumonia and rampant bacterial growth. Given the likely role of the hyperinflammatory response in accelerating the destruction of tissue, in this work we evaluated the therapeutic potential of the inducible cytoprotective enzyme heme oxygenase 1 (HO-1) against primary pneumonic plague. On its own, the HO-1 inducer cobalt protoporphyrin IX (CoPP) provided mice protection from lethal challenge with Y. pestis CO92 with improved pulmonary bacterial clearance and a dampened inflammatory response compared to vehicle-treated mice. Furthermore, CoPP treatment combined with doxycycline strongly enhanced protection in a rat aerosol challenge model. Compared to doxycycline alone, CoPP treatment increased survival, with a 3-log decrease in median bacterial titer recovered from the lungs and the general absence of a systemic hyperinflammatory response. In contrast, treatment with the HO-1 inhibitor SnPP had no detectable impact on doxycycline efficacy. The combined data indicate that countering inflammatory toxicity by therapeutically inducing HO-1 is effective in reducing the rampant growth of Y. pestis and preventing pneumonic plague.

DISCUSSION

Pneumonic plague is a deadly disease that consists of fulminant bronchopneumonia and severe sepsis. In this work, we showed that this can be prevented by treatment with the heme analog and inducer of HO-1 expression, cobalt protoporphyrin IX. On its own, CoPP treatment of mice appeared to reduce inflammatory toxicity, rather than suppress cytokine production during pulmonary Y. pestis infection. This allowed for improved bacterial clearance by the innate immune response. Synergistic protection with antibiotics was observed in a rat doxycycline treatment model. Since it is known that doxycycline efficacy is dependent on host neutrophils, these data suggest that HO-1 may improve the neutrophilic response to Y. pestis (40).

Previous work has established a role for HO-1 in improving the bactericidal mechanisms of neutrophils, and in decreasing damage to tissues caused by release of reactive oxygen species (ROS) by neutrophils (41, 42). In oxygen-rich environments, such as the lungs, free iron generated as a result of hemolysis leads to the generation of ROS that is proinflammatory and cytotoxic to cells (21). Highly virulent and invasive pathogens, such as Y. pestis, are likely able to exploit this response and grow, resulting in a feedback loop of neutrophilic inflammation and tissue damage that favors bacterial growth. Further protection from ROS may be provided by biliverdin and CO, produced by heme degradation, which have antiapoptotic and anti-inflammatory effects that could dampen immunopathology (22). Future work examining the activity of CoPP-treated macrophages or neutrophils in vitro and in vivo should be informative in understanding which, if any, of these mechanisms results in protection from pneumonic plague.

CoPP allows for Nrf2-regulated gene expression, an anti-oxidant program with pleiotropic effects, including an overall suppression of the inflammatory response (26). During Y. pestis infection of mice, however, this response was not observed, and in fact, increased IL-6 was found. This may be a consequence of modulation of host cell signaling by Y. pestis virulence factors or is an indication that essential costimulatory signals were not present. Without doxycycline, CoPP provided moderate protection and, in fact, it appeared that loss of protection may have been caused by off-target effects. For example, we found abnormally low ALP, elevated IL-6, and other modest changes in the serum of mice in the CoPP treatment group that may suggest liver toxicity. Other heme-binding proteins, primarily cytochrome P450 in the liver, are known to bind to and be inhibited by CoPP (43). Alternatively, overproduction of HO-1 in the liver may have unwanted effects (44). Additional investigation is needed to understand the mechanism underlying these observations, whether it is caused by HO-1 or CoPP directly, and whether reducing this effect improves protection. Nevertheless, the targeting of HO-1 or another cytoprotective mechanism to limit inflammatory damage is a promising treatment strategy for pneumonic plague.

Here, we observed an unexpected difference between male and female SD rats in their susceptibility to aerosol challenge with Y. pestis. In human plague, there are no known sex dependent differences in susceptibility, though historically, there have been more male than female plague victims, Sexual dimorphism has been frequently documented in infectious diseases, and in general, females show greater humoral and cell mediated responses than males of the same age and species, making females more resistant (45). In contrast, in the pneumonic plague model, SD rat females were more sensitive to infection by aerosolized Y. pestis, suggesting the effects we observed may result from the unique host-pathogen interactions that define plague. Since the disease course appeared to have the same kinetics and outcome in both sexes, we think it likely that the difference in susceptibility relates to an early event that impacts the initiation of infection. In an endotoxin challenge model, the inflammatory response of the SD rat female involved higher production of inflammatory cytokines from alveolar macrophages (46). Alveolar macrophages are early cellular targets of Y. pestis infection of the lungs, where such an effect could impact the initiation of infection (47). Future treatment and vaccine studies of plague should include male and female animals until there is a better understanding of the mechanism underlying sex-dependent susceptibility in the SD rat.

Malariaplasmodin kuparin tarve

• Format: Abstract

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Malar J. 2012 Nov 29;11:397. doi: 10.1186/1475-2875-11-397.
A Plasmodium falciparum copper-binding membrane protein with copper transport motifs.
Choveaux DL¹, Przyborski JM, Goldring JP.
Abstract

BACKGROUND:
Copper is an essential catalytic co-factor for metabolically important cellular enzymes, such as cytochrome-c oxidase. Eukaryotic cells acquire copper through a copper transport protein and distribute intracellular copper using molecular chaperones. The copper chelator, neocuproine, inhibits Plasmodium falciparum ring-to-trophozoite transition in vitro, indicating a copper requirement for malaria parasite development. How the malaria parasite acquires or secretes copper still remains to be fully elucidated.
METHODS:
PlasmoDB was searched for sequences corresponding to candidate P. falciparum copper-requiring proteins. The amino terminal domain of a putative P. falciparum copper transport protein was cloned and expressed as a maltose binding fusion protein. The copper binding ability of this protein was examined. Copper transport protein-specific anti-peptide antibodies were generated in chickens and used to establish native protein localization in P. falciparum parasites by immunofluorescence microscopy.
RESULTS:
Six P. falciparum copper-requiring protein orthologs and a candidate P. falciparum copper transport protein (PF14_0369), containing characteristic copper transport protein features, were identified in PlasmoDB. The recombinant amino terminal domain of the transport protein bound reduced copper in vitro and within Escherichia coli cells during recombinant expression. Immunolocalization studies tracked the copper binding protein translocating from the erythrocyte plasma membrane in early ring stage to a parasite membrane as the parasites developed to schizonts. The protein appears to be a PEXEL-negative membrane protein.
CONCLUSION:
Plasmodium falciparum parasites express a native protein with copper transporter characteristics that binds copper in vitro. Localization of the protein to the erythrocyte and parasite plasma membranes could provide a mechanism for the delivery of novel anti-malarial compounds.
PMID:
23190769
PMCID:
PMC3528452
DOI:
10.1186/1475-2875-11-397
[Indexed for MEDLINE]
Free PMC Article
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Tuberkuloosibakteerin entsyymi MtADPRaasi

https://link.springer.com/article/10.1007%2Fs10863-016-9681-9

• Published: 28 September 2016

Kinetic and mutational studies of the adenosine diphosphate ribose hydrolase from Mycobacterium tuberculosis

• Suzanne F. O’Handley,
• Puchong Thirawatananond,
• Lin-Woo Kang,
• Jennifer E. Cunningham,
• J. Alfonso Leyva,
• L. Mario Amzel &
• Sandra B. Gabelli 

Journal of Bioenergetics and Biomembranes volume 48, pages557–567(2016)Cite this article

• 249 Accesses
• 2 Citations
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Abstract

Mycobacterium tuberculosis represents one of the world’s most devastating infectious agents – with one third of the world’s population infected and 1.5 million people dying each year from this deadly pathogen. As part of an effort to identify targets for therapeutic intervention, we carried out the kinetic characterization of the product of gene rv1700 of M. tuberculosis. Based on its sequence and its structure, the protein had been tentatively identified as a pyrophosphohydrolase specific for adenosine diphosphate ribose (ADPR), a compound involved in various pathways including oxidative stress response and tellurite resistance. In this work we carry out a kinetic, mutational and structural investigation of the enzyme, which provides a full characterization of this Mt-ADPRase. Optimal catalytic rates were achieved at alkaline pH (7.5–8.5) with either 0.5–1 mM Mg²⁺ or 0.02–1 mM Mn²⁺. K _m and k _cat values for hydrolysis of ADPR with Mg²⁺ ions are 200 ± 19 μM and 14.4 ± 0.4 s⁻¹, and with Mn²⁺ ions are 554 ± 64 μM and 28.9 ± 1.4 s⁻¹. Four residues proposed to be important in the catalytic mechanism of the enzyme were individually mutated and the kinetics of the mutant enzymes were characterized. In the four cases, the K _m increased only slightly (2- to 3-fold) but the k _cat decreased significantly (300- to 1900-fold), confirming the participation of these residues in catalysis. An analysis of the sequence and structure conservation patterns in Nudix ADPRases permits an unambiguous identification of members of the family and provides insight into residues involved in catalysis and their participation in substrate recognition in the Mt-ADPRase.

Trypanosoman makrofomeeni

Trypanosoma crusei
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4827093/

Sci Rep. 2016; 6: 24213.

Published online 2016 Apr 11. doi: 10.1038/srep24213

PMCID: PMC4827093

PMID: 27064071

Proximal ADP-ribose Hydrolysis in Trypanosomatids is Catalyzed by a Macrodomain

Teemu Haikarainen¹ and Lari Lehtiö^a,1

Abstract

ADP-ribosylation is a ubiquitous protein modification utilized by both prokaryotes and eukaryotes for several cellular functions, such as DNA repair, proliferation and cell signaling. Higher eukaryotes, such as humans, utilize various enzymes to reverse the modification and to regulate ADP-ribose dependent signaling. In contrast, some lower eukaryotes, including trypanosomatids, lack many of these enzymes and therefore have a much more simplified ADP-ribose metabolism. Here we identified and characterized ADP-ribose hydrolases from Trypanosoma brucei and Trypanosoma cruzi, which are homologous to human O-acetyl-ADP-ribose deacetylases MacroD1 and MacroD2. The enzymes are capable for hydrolysis of protein linked ADP-ribose and a product of sirtuin-mediated lysine deacetylation, O-acetyl-ADP-ribose. Crystal structures of the trypanosomatid macrodomains revealed a conserved catalytic site with distinct differences to human MacroD1 and MacroD2.

 ADP-ribosylation 
is a covalent modification where one (mono) or multiple  (poly) ADP-ribose units are attached to a target protein. In eukaryotes, the modification is catalyzed by poly-ADP-ribose polymerases (PARPs), silent information regulators (sirtuins) and poorly characterized membrane anchored arginine ADP-ribosyltransferases (ARTs)^1,2,3.

 ADP-ribosylation regulates several cellular events, including DNA repair, cell cycle progression, transcription and cell death. In humans over 20 enzymes have been found to catalyze ADP-ribosylation and modification can be reversed by various enzymes:
 PARG (poly(ADP-ribose) glycohydrolase) and
 ARH3 (ADP-ribosylhydrolase 3), which can hydrolyze ADP-ribose polymer leaving the proximal ADP-ribose molecule 8mono-ADP-ribosyl)attached to the modified protein;
MacroD1 and MacroD2, which cleave the proximal mono-ADP-ribosyl;
 and OARD1 (O-acetyl-ADP-ribose deacetylase 1), which is capable of hydroxylation of the proximal mono-ADP-ribosylation and also cleaving the entire PAR en bloc⁴.
There are also less characterized enzymes involved in ADP-ribose hydrolysis, namely ARH1, which hydrolyzes mono-ADP-ribosylated arginines⁵ and Nudix hydrolase 16, which is able to remove both mono-and poly-ADP-ribosylation leaving the modified protein with a ribose-5′-phosphate⁶. MacroD1, MacroD2, OARD1 and ARH3 have also O-acetyl-ADP-ribose hydrolysis activity as they are capable of removing the acetyl group from O-acetyl-ADP-ribose produced by sirtuin-mediated lysine deacetylation^7,8,9.


( Merkitystä trypanosomassa:)
 Trypanosoma brucei and Trypanosoma cruzi are parasitic protozoa responsible for severe human and animal diseases. T. brucei is the causative agent of African trypanosomiasis or sleeping sickness and T. cruzi is responsible for South American trypanosomiasis or Chagas Disease.
 These parasites seem to have a highly simplified ADP-ribose metabolism compared to higher eukaryotes, such as humans.
They contain a single PARP opposed to 17 PARPs found in humans^10,11,12 and have two (T. cruzi) or three (T. brucei) sirtuins, while humans have seven (although not all have ADP-ribosylating activity)¹³.
Only one ADP-ribose hydrolase, PARG, has been characterized from the parasites¹⁴.

 Recently, a phylogenetic analysis of proteins linked to ADP-ribose metabolism identified a single MacroD1/MacroD2 homologue in T. brucei¹⁵. We analyzed the available genome sequences and observed that besides PARG and MacroD1/MacroD2 homologue, T. brucei and T. cruzi do not have other known enzymes capable for the hydrolysis of ADP-ribose. We show that the trypanosomatid MacroD1/MacroD2 homologues are hydrolyzing both protein-linked ADP-ribose and free O-acetyl-ADP-ribose (O-AADPR) The crystal structures of the enzymes reveal highly conserved macrodomain folds containing conserved ADP-ribose binding sites.

Results

Identification and characterization of trypanosomal proximal ADP-ribose hydrolases

ADP-ribose hydrolyzing enzymes of T. cruzi and T. brucei were searched from NCBI non-redundant database using known ADP-ribose hydrolases from human (PARG, ARH3, ARH1, OARD1, MacroD1, MacroD2 and Nudix hydrolase 16) as a query (Fig. 1A).
 Only one putative ADP-ribose hydrolase was identified from the parasites, which had homology to MacroD1 and MacroD2 (Fig. 1B). We named these proteins as Trypanosoma brucei MacroD-like protein (TbMDO) and Trypanosoma cruzi MacroD-like protein (TcMDO). It should be noted that T. brucei and T. cruzi contain putative Nudix hydrolases but they have very low identities to human Nudix hydrolase 16 and are more similar to other human Nudix proteins.
Human MacroD1 and MacroD2 function as ADP-ribose hydrolases cleaving ADP-ribose from target proteins, as well as O-acetyl-ADP-ribose deacetylases hydrolyzing O-acetyl-ADP-ribose produced by sirtuins during lysine deacetylation (Fig. 2A).

Trypanosomiaasi

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3831158/