Publications by authors named "Haytham M Wahba"

7 Publications

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A hydride transfer complex reprograms NAD metabolism and bypasses senescence.

Mol Cell 2021 09;81(18):3848-3865.e19

CRCHUM, 900 Saint-Denis St, Montréal, QC H2X 0A9, Canada; Département de Biochimie et Médecine Moléculaire, Université de Montréal, Montréal, QC H3C 3J7, Canada. Electronic address:

Metabolic rewiring and redox balance play pivotal roles in cancer. Cellular senescence is a barrier for tumorigenesis circumvented in cancer cells by poorly understood mechanisms. We report a multi-enzymatic complex that reprograms NAD metabolism by transferring reducing equivalents from NADH to NADP. This hydride transfer complex (HTC) is assembled by malate dehydrogenase 1, malic enzyme 1, and cytosolic pyruvate carboxylase. HTC is found in phase-separated bodies in the cytosol of cancer or hypoxic cells and can be assembled in vitro with recombinant proteins. HTC is repressed in senescent cells but induced by p53 inactivation. HTC enzymes are highly expressed in mouse and human prostate cancer models, and their inactivation triggers senescence. Exogenous expression of HTC is sufficient to bypass senescence, rescue cells from complex I inhibitors, and cooperate with oncogenic RAS to transform primary cells. Altogether, we provide evidence for a new multi-enzymatic complex that reprograms metabolism and overcomes cellular senescence.
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http://dx.doi.org/10.1016/j.molcel.2021.08.028DOI Listing
September 2021

Characterization of a C-Terminal SUMO-Interacting Motif Present in Select PIAS-Family Proteins.

Structure 2020 05 28;28(5):573-585.e5. Epub 2020 Apr 28.

Département de Biochimie et Médicine Moléculaire, Université de Montréal, C.P. 6128 Succursale Centre-Ville, Montréal, QC H3C 3J7, Canada. Electronic address:

The human PIAS proteins are small ubiquitin-like modifier (SUMO) E3 ligases that participate in important cellular functions. Several of these functions depend on a conserved SUMO-interacting motif (SIM) located in the central region of all PIAS proteins (SIM1). Recently, it was determined that Siz2, a yeast homolog of PIAS proteins, possesses a second SIM at its C terminus (SIM2). Sequence alignment indicates that a SIM2 is also present in PIAS1-3, but not PIAS4. Using biochemical and structural studies, we demonstrate PIAS-SIM2 binds to SUMO1, but that phosphorylation of the PIAS-SIM2 or acetylation of SUMO1 alter this interaction in a manner distinct from what is observed for the PIAS-SIM1. We also show that the PIAS-SIM2 plays a key role in formation of a UBC9-PIAS1-SUMO1 complex. These results provide insights into how post-translational modifications selectively regulate the specificity of multiple SIMs found in the PIAS proteins by exploiting the plasticity built into the SUMO-SIM binding interface.
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http://dx.doi.org/10.1016/j.str.2020.04.002DOI Listing
May 2020

Acetylation of SUMO1 Alters Interactions with the SIMs of PML and Daxx in a Protein-Specific Manner.

Structure 2020 02 23;28(2):157-168.e5. Epub 2019 Dec 23.

Département de Biochimie et Médicine Moléculaire, Université de Montréal, C.P. 6128 Succursale Centre-Ville, Montréal, QC H3C 3J7, Canada. Electronic address:

The interactions between SUMO proteins and SUMO-interacting motif (SIM) in nuclear bodies formed by the promyelocytic leukemia (PML) protein (PML-NBs) have been shown to be modulated by either phosphorylation of the SIMs or acetylation of SUMO proteins. However, little is known about how this occurs at the atomic level. In this work, we examined the role that acetylation of SUMO1 plays on its binding to the phosphorylated SIMs (phosphoSIMs) of PML and Daxx. Our results demonstrate that SUMO1 binding to the phosphoSIM of either PML or Daxx is dramatically reduced by acetylation at either K39 or K46. However, acetylation at K37 only impacts binding to Daxx. Structures of acetylated SUMO1 variants bound to the phosphoSIMs of PML and Daxx demonstrate that there is structural plasticity in SUMO-SIM interactions. The plasticity observed in these structures provides a robust mechanism for regulating SUMO-SIM interactions in PML-NBs using signaling generated post-translational modifications.
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http://dx.doi.org/10.1016/j.str.2019.11.019DOI Listing
February 2020

Structural and Biochemical Characterization of Organotin and Organolead Compounds Binding to the Organomercurial Lyase MerB Provide New Insights into Its Mechanism of Carbon-Metal Bond Cleavage.

J Am Chem Soc 2017 01 3;139(2):910-921. Epub 2017 Jan 3.

Département de Biochimie et Médicine Moléculaire, Université de Montréal , Montréal, Quebec H3C 3J7 Canada.

The organomercurial lyase MerB has the unique ability to cleave carbon-Hg bonds, and structural studies indicate that three residues in the active site (C96, D99, and C159 in E. coli MerB) play important roles in the carbon-Hg bond cleavage. However, the role of each residue in carbon-metal bond cleavage has not been well-defined. To do so, we have structurally and biophysically characterized the interaction of MerB with a series of organotin and organolead compounds. Studies with two known inhibitors of MerB, dimethyltin (DMT) and triethyltin (TET), reveal that they inhibit by different mechanisms. In both cases the initial binding is to D99, but DMT subsequently binds to C96, which induces a conformation change in the active site. In contrast, diethyltin (DET) is a substrate for MerB and the Sn product remains bound in the active site in a coordination similar to that of Hg following cleavage of organomercurial compounds. The results with analogous organolead compounds are similar in that trimethyllead (TML) is not cleaved and binds only to D99, whereas diethyllead (DEL) is a substrate and the Pb product remains bound in the active site. Binding and cleavage is an exothermic reaction, while binding to D99 has negligible net heat flow. These results show that initial binding of organometallic compounds to MerB occurs at D99 followed, in some cases, by cleavage and loss of the organic moieties and binding of the metal ion product to C96, D99, and C159. The N-terminus of MerA is able to extract the bound Pb but not the bound Sn. These results suggest that MerB could be utilized for bioremediation applications, but certain organolead and organotin compounds may present an obstacle by inhibiting the enzyme.
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http://dx.doi.org/10.1021/jacs.6b11327DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6040655PMC
January 2017

Structural and Biochemical Characterization of a Copper-Binding Mutant of the Organomercurial Lyase MerB: Insight into the Key Role of the Active Site Aspartic Acid in Hg-Carbon Bond Cleavage and Metal Binding Specificity.

Biochemistry 2016 Feb 11;55(7):1070-81. Epub 2016 Feb 11.

Department of Chemistry, Dartmouth College , Hanover, New Hampshire 03755, United States.

In bacterial resistance to mercury, the organomercurial lyase (MerB) plays a key role in the detoxification pathway through its ability to cleave Hg-carbon bonds. Two cysteines (C96 and C159; Escherichia coli MerB numbering) and an aspartic acid (D99) have been identified as the key catalytic residues, and these three residues are conserved in all but four known MerB variants, where the aspartic acid is replaced with a serine. To understand the role of the active site serine, we characterized the structure and metal binding properties of an E. coli MerB mutant with a serine substituted for D99 (MerB D99S) as well as one of the native MerB variants containing a serine residue in the active site (Bacillus megaterium MerB2). Surprisingly, the MerB D99S protein copurified with a bound metal that was determined to be Cu(II) from UV-vis absorption, inductively coupled plasma mass spectrometry, nuclear magnetic resonance, and electron paramagnetic resonance studies. X-ray structural studies revealed that the Cu(II) is bound to the active site cysteine residues of MerB D99S, but that it is displaced following the addition of either an organomercurial substrate or an ionic mercury product. In contrast, the B. megaterium MerB2 protein does not copurify with copper, but the structure of the B. megaterium MerB2-Hg complex is highly similar to the structure of the MerB D99S-Hg complexes. These results demonstrate that the active site aspartic acid is crucial for both the enzymatic activity and metal binding specificity of MerB proteins and suggest a possible functional relationship between MerB and its only known structural homologue, the copper-binding protein NosL.
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http://dx.doi.org/10.1021/acs.biochem.5b01298DOI Listing
February 2016

Antimicrobial activity of some Clerodendrum species from Egypt.

Nat Prod Res 2013 17;27(11):1032-6. Epub 2012 May 17.

Pharmacognosy Department, Faculty of Pharmacy, Beni-Suef University, Beni-Suef, Egypt.

Chloroformic and methanolic extracts of four Clerodendrum species cultivated in Egypt were screened for antimicrobial activities. Chloroformic extracts of the flowers of Clerodendrum chinense and Clerodendrum splendens were active against Plasmodium falciparum (IC50 < 10 µg mL(-1)). Chloroformic extracts of the stem and flowers of C. chinense were active against Trypanosoma cruzi (IC50 = 1.21 and 1.12 µg mL(-1), respectively) with marginal cytotoxicity. Chloroformic extracts of the leaves of C. chinense and C. splendens showed promising activities against T. cruzi (IC50 = 3.39 and 1.98 µg mL(-1), respectively) without cytotoxic effect on a human cell line. None of the selected plants showed significant activity against Gram-negative or Gram-positive bacteria or Candida albicans. Verbascoside, a phenyl propanoid glycoside isolated from the leaves of C. chinense, showed marginal activity against T. cruzi. Rengyolone, a cyclohexyl ethanoid isolated from the leaves of C. chinense, showed a broad but not specific activity against the tested organisms.
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http://dx.doi.org/10.1080/14786419.2012.686905DOI Listing
January 2014

Chemical and biological investigation of some Clerodendrum species cultivated in Egypt.

Pharm Biol 2011 Jan 26;49(1):66-72. Epub 2010 Aug 26.

Pharmacognosy Department, Faculty of Pharmacy, Beni-Sueif University, Beni-Sueif, Egypt.

Context: Phytochemical investigation of Clerodendrum chinense (Osbeck) Mabberley (Lamiaceae) cultivated in Egypt and evaluation for anti-inflammatory, analgesic, and antipyretic effects of the methanol and chloroform extracts of Clerodendrum chinense, Clerodendrum indicum (L.) Kuntze, Clerodendrum glabrum E. Meyer.

Objective: The objective of this study was to investigate the anti-inflammatory, analgesic, and antipyretic effects of the methanol and chloroform extracts of Clerodendrum species under investigation.

Materials And Methods: Air-dried powdered leaves of C. chinense were extracted with MeOH 80%. This extract was fractionated with successive portions of chloroform, ethyl acetate and n-butanol. By further fractionation through silica gel, polyamide and reversed phase column chromatography several compounds were isolated which were elucidated by nuclear magnetic resonance (NMR) and mass spectroscopy. For biological study, the powdered leaves of C. chinense, C. indicum and C. glabrum were extracted by chloroform and then extracted with methanol. The acute anti-inflammatory effect of tested extracts of the leaves of Clerodendrum species under investigation was estimated by carrageenan-induced rat paw edema. Antipyretic effect was evaluated and compared with that of paracetamol as standard using the yeast-induced hyperthermia method on female albino rats. Analgesic effect was evaluated and compared with that of Novalgin (metamizol sodium) as standard using an electric current anxious stimulus.

Results: Verbascoside, isoverbascoside, decaffeoylverbascoside, hispidulin, lupeol and icariside B5 were isolated from the leaves of C. chinense for the first time. Cornoside and rengyolone were also isolated. The methanol extract of the leaves of C. chinense and verbascoside showed significant analgesic, anti-inflammatory and antipyretic effects.

Conclusion: The present study provided a scientific validation of the traditional claims suggested for C. chinense.
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http://dx.doi.org/10.3109/13880209.2010.494674DOI Listing
January 2011
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