Metal effect on intein splicing: A review

Panda, Sunita; Nanda, Ananya; Nasker, Sourya Subhra; Sen, Debjani; Mehra, Ashwaria; Nayak, Sasmita

doi:10.1016/j.biochi.2021.03.006

Cited by 10 publications

(6 citation statements)

References 102 publications

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“…61 Other metal ions, such as Cd 2+ , Ni 2+ , and Co 2+ , have shown weaker inhibition than Zn 2+ on splicing of the RecA intein from Mycobacterium tuberculosis (Mtb) and naturally split DnaE intein from Synechocystis sp. 62,63 Two reactive chlorine species, hypochlorous acid and N-chlorotaurine, can reversibly block splicing of the DnaB inteins from Mycobacterium leprae and Msm in vitro. 64 Platinum-containing compounds such as cisplatin inhibit splicing of the RecA intein of Mtb with an IC 50 of 2 μM in vitro.…”

supporting

confidence: 62%

Calcimycin Inhibits Cryptococcus neoformans In Vitro and In Vivo by Targeting the Prp8 Intein Splicing

Tharappel

Zhu

et al. 2022

ACS Infect. Dis.

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Drug resistance is a significant concern in the treatment of diseases, including cryptococcosis caused by Cryptococcus neoformans (Cne) and Cryptococcus gattii (Cga). Alternative drug targets are necessary to overcome drug resistance before it attains a critical stage. Splicing of inteins from pro-protein precursors is crucial for activities of essential proteins hosting intein elements in many organisms, including human pathogens such as Cne and Cga. Through a high-throughput screening, we identified calcimycin (CMN) as a potent Prp8 intein splicing inhibitor with a minimum inhibitory concentration (MIC) of 1.5 μg/mL against the wild-type Cne-H99 (Cne-WT or Cne). In contrast, CMN inhibited the intein-less mutant strain (Cne-Mut) with a 16-fold higher MIC. Interestingly, Aspergillus fumigatus and a few Candida species were resistant to CMN. Further studies indicated that CMN reduced virulence factors such as urease activity, melanin production, and biofilm formation in Cne. CMN also inhibited Cne intracellular infection in macrophages. In a target-specific split nanoluciferase assay, the IC50 of CMN was 4.6 μg/mL. Binding of CMN to recombinant Prp8 intein was demonstrated by thermal shift assay and microscale thermophoresis. Treating Cne cells with CMN reduced intein splicing. CMN was fungistatic and showed a synergistic effect with the known antifungal drug amphotericin B. Finally, CMN treatment at 20 mg/kg body weight led to 60% reduction in lung fungal load in a cryptococcal pulmonary infection mouse model. Overall, CMN represents a potent antifungal with a novel mechanism of action to treat Cne and possibly Cga infections.

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supporting

confidence: 62%

Calcimycin Inhibits Cryptococcus neoformans In Vitro and In Vivo by Targeting the Prp8 Intein Splicing

Tharappel

Zhu

et al. 2022

ACS Infect. Dis.

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“…The DnaBi1 of M. smegmatis responds to oxidative stress, whereas DnaBi2 is not ( Kelley et al, 2018 ). Cd 2+ , Ni 2+ and Co 2+ have also shown inhibitory activity, although weaker than Zn 2+ , in splicing of the RecA intein from M. tuberculosis and of naturally split DnaE intein from Synechocystis sp., whereas there is no inhibition by Mg 2+ and Ca 2+ ( Mills and Paulus, 2001 ; Panda et al, 2021 ).…”

Section: Introductionmentioning

confidence: 99%

Inteins as Drug Targets and Therapeutic Tools

Tharappel

2022

Front. Mol. Biosci.

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Multidrug-resistant pathogens are of significant concern in recent years. Hence new antifungal and anti-bacterial drug targets are urgently needed before the situation goes beyond control. Inteins are polypeptides that self-splice from exteins without the need for cofactors or external energy, resulting in joining of extein fragments. Inteins are present in many organisms, including human pathogens such as Mycobacterium tuberculosis, Cryptococcus neoformans, C. gattii, and Aspergillus fumigatus. Because intein elements are not present in human genes, they are attractive drug targets to develop antifungals and antibiotics. Thus far, a few inhibitors of intein splicing have been reported. Metal-ions such as Zn2+ and Cu2+, and platinum-containing compound cisplatin inhibit intein splicing in M. tuberculosis and C. neoformans by binding to the active site cysteines. A small-molecule inhibitor 6G-318S and its derivative 6G-319S are found to inhibit intein splicing in C. neoformans and C. gattii with a MIC in nanomolar concentrations. Inteins have also been used in many other applications. Intein can be used in activating a protein inside a cell using small molecules. Moreover, split intein can be used to deliver large genes in experimental gene therapy and to kill selected species in a mixed population of microbes by taking advantage of the toxin-antitoxin system. Furthermore, split inteins are used in synthesizing cyclic peptides and in developing cell culture model to study infectious viruses including SARS-CoV-2 in the biosafety level (BSL) 2 facility. This mini-review discusses the recent research developments of inteins in drug discovery and therapeutic research.

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“…This post-translational auto-excision of the intervening intein protein is critical for the formation of an active protein [1][2][3][4]. Splicing and cleavage products (Figure 1C) are obtained through a series of nucleophilic displacement reactions mediated in a coordinated fashion by the catalytic residues [5][6][7][8][9]. Control over this reversible interruption of the functional form of the host exteins can play a regulatory role in protein activation [10,11].…”

Section: Introductionmentioning

confidence: 99%

SufB intein splicing in Mycobacterium tuberculosis is influenced by two remote conserved N-extein histidines

et al. 2022

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Inteins are auto-processing domains that implement a multistep biochemical reaction termed protein splicing, marked by cleavage and formation of peptide bonds. They excise from a precursor protein, generating a functional protein via covalent bonding of flanking exteins. We report the kinetic study of splicing and cleavage reaction in [Fe–S] cluster assembly protein SufB from Mycobacterium tuberculosis (Mtu). Although it follows a canonical intein splicing pathway, distinct features are added by extein residues present in the active site. Sequence analysis identified two conserved histidines in the N-extein region; His-5 and His-38. Kinetic analyses of His-5Ala and His-38Ala SufB mutants exhibited significant reductions in splicing and cleavage rates relative to the SufB wildtype (WT) precursor protein. Structural analysis and molecular dynamics (MD) simulations suggested that Mtu SufB displays a unique mechanism where two remote histidines work concurrently to facilitate N-terminal cleavage reaction. His-38 is stabilized by the solvent-exposed His-5, and can impact N–S acyl shift by direct interaction with the catalytic Cys1. Development of inteins as biotechnological tools or as pathogen-specific novel antimicrobial targets requires a more complete understanding of such unexpected roles of conserved extein residues in protein splicing.

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Metal effect on intein splicing: A review

Cited by 10 publications

References 102 publications

Calcimycin Inhibits Cryptococcus neoformans In Vitro and In Vivo by Targeting the Prp8 Intein Splicing

Calcimycin Inhibits Cryptococcus neoformans In Vitro and In Vivo by Targeting the Prp8 Intein Splicing

Inteins as Drug Targets and Therapeutic Tools

SufB intein splicing in Mycobacterium tuberculosis is influenced by two remote conserved N-extein histidines

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