Gregory M Cook

University of Otago · Department of Microbiology and Immunology

Transport of proteins across and into membranes is a fundamental biological process with the vast majority being conducted by the ubiquitous Sec machinery. In bacteria, this is usually achieved when the SecY-complex engages the cytosolic ATPase SecA (secretion) or translating ribosomes (insertion). Great strides have been made towards understanding...

Oxidation of malate to oxaloacetate, catalyzed by either malate dehydrogenase (Mdh) or malate quinone oxidoreductase (Mqo), is a critical step of the tricarboxylic acid (TCA) cycle. Both Mqo and Mdh are found in most bacterial genomes, but the level of functional redundancy between these enzymes remains unclear. A bioinformatic survey revealed that...

Increasing antimicrobial resistance compels the search for next-generation inhibitors with differing or multiple molecular targets. In this regard, energy conservation in Mycobacterium tuberculosis has been clinically validated as a promising new drug target for combatting drug-resistant strains of M. tuberculosis. Here, we show that HM2-16F, a 6-s...

Objectives There is an urgent need for novel drugs that target unique cellular pathways to combat infections caused by Mycobacterium tuberculosis. CRISPR interference (CRISPRi)-mediated transcriptional repression has recently been developed for use in mycobacteria as a genetic tool for identifying and validating essential genes as novel drug target...

Multidrug resistance is a major threat to global elimination of tuberculosis (TB). We performed phenotypic drug-susceptibility testing and whole-genome sequencing for 309 isolates from 342 consecutive patients who were given a diagnosis of TB in Yangon, Myanmar, during July 2016‒June 2018. We identified isolates by using the GeneXpert platform to e...

Disruption of redox homeostasis in mycobacteria causes irreversible stress induction and cell death. Here, we report the dioxonaphthoimidazolium scaffold as a novel redox cycling antituberculosis chemotype with potent bactericidal activity against growing and nutrient-starved phenotypically drug-resistant nongrowing bacteria. Maximal potency was de...

Mycobacterium tuberculosis remains a leading cause of death for which new drugs are needed. The identification of drug targets has been advanced by high‐throughput and targeted genetic deletion strategies. Each though has limitations including the inability to distinguish between levels of vulnerability, lethality and scalability as a molecular too...

Mycobacterium tuberculosis remains a leading cause of infectious disease morbidity and mortality for which new drug combination therapies are needed. Combinations of respiratory inhibitors can have synergistic or synthetic lethal interactions suggesting that regimens with multiple bioenergetic inhibitors will drastically shorten treatment times. Ho...

New drugs are urgently needed to combat the global TB epidemic. Targeting simultaneously multiple respiratory enzyme complexes of Mycobacterium tuberculosis is regarded as one of the most effective treatment options to shorten drug administration regimes, and reduce the opportunity for the emergence of drug resistance. During infection and prolifer...

With the emergence of multidrug-resistant strains of Mycobacterium tuberculosis (MDR-TB) and extensive drug-resistant strains (XDR-TB), there is an urgent need to develop novel drugs for the treatment of tuberculosis. Here, we designed and synthesized a series of 5-methylpyrimidopyridone analogues as potential antitubercular agents. The most potent...

Malacidin A is a novel calcium-dependent lipopeptide antibiotic with excellent activity against Gram-positive pathogens. Herein, a concise and robust synthetic route toward malacidin A is reported, employing 9-fluorenylmethoxycarbonyl solid-phase peptide synthesis of a linear precursor, including late-stage incorporation of the lipid tail, followed...

Methane emissions from enteric fermentation in the ruminant digestive system generated by methanogenic archaea are a significant contributor to anthropogenic greenhouse gas emissions. Additionally, methane produced as an end-product of enteric fermentation is an energy loss from digested feed. To control the methane emissions from ruminants, extens...

Multidrug resistant (MDR) tuberculosis (TB) is defined by the resistance of Mycobacterium tuberculosis , the causative organism, to the first-line antibiotics rifampicin and isoniazid. Mitigating or reversing resistance to these drugs offers a means of preserving and extending their use in TB treatment. R-loops are RNA/DNA hybrids that are formed i...

Succinate is a major focal point in mycobacterial metabolism and respiration, serving as both an intermediate of the TCA cycle and a direct electron donor for the respiratory chain. Mycobacterium tuberculosis encodes multiple enzymes predicted to be capable of catalyzing the oxidation of succinate to fumarate, including two different succinate dehy...

Neutrophils are often the major leukocyte at sites of mycobacterial infection, yet little is known about their ability to kill mycobacteria. In this study we have investigated whether the potent antibacterial oxidant hypochlorous acid (HOCl) contributes to killing of Mycobacterium smegmatis when this bacterium is phagocytosed by human neutrophils....

The K⁺-sparing diuretic amiloride elicits anticancer activities in multiple animal models. During our recent medicinal chemistry campaign aiming to identify amiloride analogs with improved properties for potential use in cancer, we discovered novel 6-(hetero)aryl-substituted amiloride and 5-(N,N-hexamethylene)amiloride (HMA) analogs with up to 100-...

Members of the genus Methylacidiphilum, a clade of metabolically flexible thermoacidophilic methanotrophs from the phylum Verrucomicrobia, can utilize a variety of substrates including methane, methanol, and hydrogen for growth. However, despite sequentially oxidizing methane to carbon dioxide via methanol and formate intermediates, growth on forma...

The approval of bedaquiline has placed energy metabolism in the limelight as an attractive target space for tuberculosis antibiotic development. While bedaquiline inhibits the mycobacterial F 1 F 0 ATP synthase, small molecules targeting other components of the oxidative phosphorylation pathway have been identified. Of particular interest is Telace...

The emergence of polymyxin resistance in carbapenem-resistant and extended-spectrum -lactamase (ESBL)-producing bacteria is a critical threat to human health, and alternative treatment strategies are urgently required. We investigated the ability of the hydroxyquinoline analog ionophore PBT2 to restore antibiotic sensitivity in polymyxin-resistant...

The aerobic thermoalkaliphile Caldalkalibacillus thermarum strain TA2.A1 is a member of a separate order of alkaliphilic bacteria closely related to the Bacillales order. Efforts to relate the genomic information of this evolutionary ancient organism to environmental adaptation have been thwarted by the inability to construct a complete genome. The...

A series of C-2 derivatized 8-sulfonamidoquinolines were evaluated for their antibacterial activity against the common mastitis causative pathogens Streptococcus uberis, Staphylococcus aureus and Escherichia coli, both in the presence and absence of supplementary zinc (50 µM ZnSO4). The vast majority of compounds tested were demonstrated to be sign...

Targeting energy metabolism in Mycobacterium tuberculosis (Mtb) is a new paradigm in the search for innovative anti-TB drugs. NADH:menaquinone oxidoreductase is a non-proton translocating type II NADH dehydrogenase (NDH-2) that is an essential enzyme in the respiratory chain of Mtb and is not found in mammalian mitochondria. Phenothiazines (PTZs) r...

The ohmyungsamycin and ecumicin natural product families are structurally related cyclic depsipeptides that display potent antimycobacterial activity. Herein the total synthesis of ohmyungsamycin A, deoxyecumicin, and ecumicin are reported, together with the first direct biological comparison of members of these natural product families against Myc...

Mycobacteria are major environmental microorganisms and cause many significant diseases, including tuberculosis. Mycobacteria make an unusual vitamin-like compound, F 420 , and use it to both persist during stress and resist antibiotic treatment. Understanding how mycobacteria make F 420 is important, as this process can be targeted to create new d...

Bedaquiline, an inhibitor of the mycobacterial ATP synthase, has revolutionized the treatment of Mycobacterium tuberculosis . Although a potent inhibitor, it is characterized by a poorly understood delayed time-dependent bactericidal activity. Here, we demonstrate that in contrast to bedaquiline, the transcriptional inhibition of the ATP synthase i...

The mycobacterial membrane protein Large 3 (MmpL3) is an inner membrane protein that transports trehalose-monomycolates, precursors for trehalose-dimycolates and mycolic acids that make up essential components of the mycobacterial outer membrane. Inhibition of MmpL3 weakens the mycobacterial cell wall and ultimately results in cell death in both in...

The formation of biofilms provides a formidable defense for many bacteria against antibiotics and host immune responses. As a consequence, biofilms are thought to be the root cause of most chronic infections, including those occurring on medical indwelling devices, endocarditis, urinary tract infections, diabetic and burn wounds and bone and joint...

Drug‐resistant strains of Mycobacterium tuberculosis are on the rise and have resulted in an urgent need to develop new tuberculosis (TB) drugs. Herein, the development of chromonyl‐pyrimidines as a new TB drug scaffold is presented. A library of eleven chromonyl‐pyrimidines was synthesised, from which lead compound 6‐((2‐amino‐6‐methylpyrimidin‐4‐...

More antimicrobials are used in food-producing animals than in humans, and the extensive use of medically important human antimicrobials poses a significant public health threat in the face of rising antimicrobial resistance. Therefore, the elimination of antimicrobial crossover between human and veterinary medicine is of great interest. Unfortunat...

A series of substituted sulfonamide bioisosteres of 8-hydroxyquinoline were evaluated for their antibacterial activity against the common mastitis causative pathogens Streptococcus uberis, Staphylococcus aureus and Escherichia coli, both in the presence and absence of supplementary zinc. Compounds 9a-e, 10a-c, 11a-e, 12 and 13 were demonstrated to...

F420 is a low-potential redox cofactor used by diverse bacteria and archaea. In mycobacteria, this cofactor has multiple roles, including adaptation to redox stress, cell wall biosynthesis, and activation of the clinical antitubercular prodrugs pretomanid and delamanid. A recent biochemical study proposed a revised biosynthesis pathway for F420 in...

The ability to respire and generate ATP is essential for the physiology, persistence and pathogenicity of Mycobacterium tuberculosis, which causes Tuberculosis. By employing a lead repurposing strategy, the malarial cytochrome bc1 inhibitor SCR0911 was tested against mycobacteria. Docking studies were carried out to reveal potential binding and to...

Our inability to predict which mutations could result in antibiotic resistance has made it difficult to rapidly identify the emergence of resistance, identify pre-existing resistant populations, and manage our use of antibiotics to effectively treat patients and prevent or slow the spread of resistance. Here we investigated the potential for resist...

Wherever thermodynamics allows, microbial life has evolved to transform and harness energy. Microbial life thus abounds in the most unexpected places, enabled by profound metabolic diversity. Within this diversity, energy is transformed primarily through variations on a few core mechanisms. Energy is further managed by the physiological processes o...

Type II NADH:quinone oxidoreductase (NDH-2) plays a crucial role in the respiratory chains of many organisms. Its absence in mammalian cells makes NDH-2 an attractive new target for developing antimicrobials and anti-protozoal agents. We established a novel bioelectrochemical platform to characterize the catalytic behavior of NDH-2 from Caldalkalib...

Mycobacterium tuberculosis (Mtb) is a globally distributed bacterial pathogen whose population structure has largely been shaped by the activities of its obligate human host. Oceania was the last major global region to be reached by Europeans and is the last region for which the dispersal and evolution of Mtb remains largely unexplored. Here, we in...

The dynamic interaction of the N- and C-terminal domains of mycobacterial F-ATP synthase subunit ε is proposed to contribute to efficient coupling of H+-translocation and ATP synthesis. Here, we investigate crosstalk between both subunit ε domains by introducing chromosomal atpC missense mutations in the C-terminal helix 2 of ε predicted to disrupt...

Background Digestive processes in the rumen lead to the release of methyl-compounds, mainly methanol and methylamines, which are used by methyltrophic methanogens to form methane, an important agricultural greenhouse gas. Methylamines are produced from plant phosphatidylcholine degradation, by choline trimethylamine lyase, while methanol comes from...

Cellular bioenergetics is an area showing promise for the development of new antimicrobials, antimalarials and cancer therapy. Enzymes involved in central carbon metabolism and energy generation are essential mediators of bacterial physiology, persistence and pathogenicity, lending themselves natural interest for drug discovery. In particular, succ...

To persist when nutrient sources are limited, aerobic soil bacteria metabolize atmospheric hydrogen (H2). This process is the primary sink in the global H2 cycle and supports the productivity of microbes in oligotrophic environments. H2-metabolizing bacteria possess [NiFe]-hydrogenases that oxidize H2 to subatmospheric concentrations. The soil sapr...

Bedaquiline is a newly developed anti-tuberculosis drug, conditionally approved by the United States Food and Drug Administration (USFDA) for treating drug-resistant tuberculosis in adults. Oral delivery of bedaquiline causes severe side effects such as increased hepatic aminotransferase levels and cardiac arrhythmias (prolongation of QT-interval)....

The oral K+-sparing diuretic amiloride shows anti-cancer side-activities in multiple rodent models. These effects appear to arise, at least in part, through moderate inhibition of the urokinase-type plasminogen activator (uPA, Ki = 2.4 µM), a pro-metastatic trypsin-like serine protease that is upregulated in many aggressive solid malignancies. In a...

A major constraint for developing new anti-tuberculosis drugs is the limited number of validated targets that allow eradication of persistent infections. Here, we uncover a vulner- able component of Mycobacterium tuberculosis (Mtb) persistence metabolism, the aspartate pathway. Rapid death of threonine and homoserine auxotrophs points to a distinct...

Aerobic soil bacteria metabolize atmospheric hydrogen (H2) to persist when nutrient sources are limited. This process is the primary sink in the global H2 cycle and supports the productivity of microbes in oligotrophic environments. To mediate this function, bacteria possess [NiFe]-hydrogenases capable of oxidising H2 to subatmospheric concentratio...

In this study we investigated the influence of oxygen availability on a phenotypic microtiter screen to identify new, natural product inhibitors of growth for the bovine mastitis-causing microorganisms; Streptococcus uberis, Staphylococcus aureus, and Escherichia coli. Mastitis is a common disease in dairy cattle worldwide and is a major cause of r...

The crystal structure of the F1-catalytic domain of the adenosine triphosphate (ATP) synthase has been determined from the pathogenic anaerobic bacterium Fusobacterium nucleatum. The enzyme can hydrolyse ATP but is partially inhibited. The structure is similar to those of the F1-ATPases from Caldalkalibacillus thermarum, which is more strongly inhi...

Farmed ruminants are the largest source of anthropogenic methane emissions globally. The methanogenic archaea responsible for these emissions use molecular hydrogen (H 2), produced during bacterial and eukaryotic carbohydrate fermentation, as their primary energy source. In this work, we used comparative genomic, metatranscriptomic and co-culture-b...

A library of thirty-two quinolinequinones (QQs) with various amine substituents at the 6- and 7-positions were synthesised efficiently and in good yields for evaluation as potential anti-tuberculosis agents. Mycobacterium tuberculosis growth inhibition assays demonstrated that QQs bearing moderate length alkyl chains (i.e. heptylphenylamino- and oc...

Teixobactin is a new antimicrobial with no known mechanisms of resistance. Understanding how resistance could develop will be crucial to the success and longevity of teixobactin as a new potent antimicrobial. Antimicrobial tolerance has been shown to facilitate the development of resistance, and we show that E. faecalis is intrinsically tolerant to...

The Mycobacterium tuberculosis complex lineage 4 (L4), also known as the “Euro-American” lineage, is the most widely dispersed of the seven human adapted lineages. L4 is comprised of ten sublineages including L4.4, which has a moderate global distribution and is the most common L4 sublineage in New Zealand. We have used a phylodynamics approach and...

Our inability to predict which mutations could result in antibiotic resistance has made it difficult to rapidly identify the emergence of resistance, identify pre-existing resistant populations and manage our use of antibiotics to effective treat patients and prevent or slow the spread of resistance. Here we investigated the potential for resistanc...

Enterococcus faecalis and Enterococcus faecium are human and animal gut commensals. Vancomycin-resistant enterococci (VRE) are important opportunistic pathogens with limited treatment options. Historically, the glycopeptide antibiotics vancomycin and avoparcin selected for the emergence of vancomycin resistance in human and animal isolates respecti...

Bacitracin is a cell wall targeting antimicrobial with clinical and agricultural applications. With the growing mismatch between antimicrobial resistance and development, it is essential we understand the molecular mechanisms of resistance in order to prioritize and generate new effective antimicrobials. BcrR is a unique membrane-bound one-componen...

The crystal structure of the F 1 -catalytic domain of the adenosine triphosphate (ATP) synthase has been determined from Mycobacterium smegmatis which hydrolyzes ATP very poorly. The structure of the α 3 β 3 -component of the catalytic domain is similar to those in active F 1 -ATPases in Escherichia coli and Geobacillus stearothermophilus . However...

There is a paucity of information on the unique components that pathogens use to form respiratory chains. It is not known why mycobacteria encode multiple succinate dehydrogenases (SDHs) to perform menaquinone‐linked succinate oxidation, a thermodynamically unfavorable reaction (ΔG° = +21 kJ/mol). In other bacteria, specific di‐heme SDHs overcome t...

: A series of 2‐ and 7‐substituted phthalazinones was synthesised and their potential as anti‐tubercular drugs assessed via Mycobacterium tuberculosis (mc26230) growth inhibition assays. All phthalazinones tested showed growth inhibitory activity (MIC < 100 μM), and those compounds containing lipophilic and electron‐withdrawing groups generally exh...

The ability to persist in the absence of growth triggered by low-oxygen levels is a critical process for the survival of mycobacterial species in many environmental niches. MSMEG_5243 (fsq), a gene of unknown function in Mycobacterium smegmatis, is up-regulated in response to hypoxia and regulated by DosRDosS/DosT, an oxygen- and redox-sensing two-...

New Zealand has a low burden of multi-drug resistant TB (MDR-TB), but with increased mobility within the population, rapid detection and treatment of MDR-TB is a priority from the public health point of view. Mycobacterium Reference Laboratory in LabPLUS, Auckland City Hospital receives referred Mycobacterium tuberculosis complex (MTBC) isolates fr...