Biology subjects

Abendroth, J.

Publications and source records attributed to Abendroth, J..

6 recordsLinked to original sources

bioRxiv2026

Structural Basis of Glycolytic Control in Trypanosoma cruzi: Insights from Enolase and PGI

Trypanosoma cruzi, the etiological agent of Chagas disease, depends on glycolysis for ATP production, rendering its glycolytic enzymes attractive targets for therapeutic development. Here, we report the high-resolution crystal structures of two essential glycolytic enzymes, glucose-6-phosphate isomerase (Tc PGI, 1.8 [A]) and enolase (Tc enolase, 2.4 [A]) and provide structural and computational analyses to support structure-based drug design. Tc PGI adopts a dimeric {beta} sandwich fold and features a parasite-specific 53-residue N-terminal extension and a unique C-terminal hook region which both distinguish it from its human ortholog. Tc enolase exhibits the conserved (/{beta}) 8 TIM barrel fold but harbors minor distinct structural deviations, including an extended 17 helix and a structured 1 region, which differentiate it from human isoforms. Both enzymes exhibited high thermal stability, consistent with adaptation to the parasites complex life cycle. Structure-based virtual screening using a scaffold with known multi-target potential identified distinct high-affinity inhibitors for each enzyme. Molecular dynamics simulations further confirmed stable enzyme-inhibitor interactions and favorable binding energetics. Collectively, these findings reveal structural signatures unique to T. cruzi glycolytic enzymes and lay the groundwork for the development of selective antiparasitic therapeutics.

biochemistry↗

bioRxiv2024

Enolase inhibitors as therapeutic leads for Naegleria fowleri infection

Infections with the pathogenic free-living amoebae Naegleria fowleri can lead to life-threatening illnesses including catastrophic primary amebic meningoencephalitis (PAM). Efficacious treatment options for these infections are lacking and the mortality rate remains >95% in the US. Glycolysis is very important for the infectious trophozoite lifecycle stage and inhibitors of glucose metabolism have been found to be toxic to the pathogen. Recently, human enolase 2 (ENO2) phosphonate inhibitors have been developed as lead agents to treat glioblastoma multiforme (GBM). These compounds, which cure GBM in a rodent model, are well-tolerated in mammals because enolase 1 (ENO1) is the predominant isoform used systemically. Here, we describe findings that demonstrate that these agents are potent inhibitors of N. fowleri ENO (NfENO) and are lethal to amoebae. In particular, (1-hydroxy-2-oxopiperidin-3-yl) phosphonic acid (HEX) was a potent enzyme inhibitor (IC50 value of 0.14 {+/-} 0.04 {micro}M) that was toxic to trophozoites (EC50 value of 0.21 {+/-} 0.02 {micro}M) while the reported CC50 was >300 {micro}M. Molecular docking simulation revealed that HEX binds strongly to the active site of NfENO with a binding affinity of -8.6 kcal/mol. Metabolomic studies of parasites treated with HEX revealed a 4.5 to 78-fold accumulation of glycolytic intermediates upstream of NfENO. Last, nasal instillation of HEX increased longevity of amoebae-infected rodents. Two days after infection, animals were treated for 10 days with 3 mg/kg HEX, followed by one week of observation. At the conclusion of the experiment, eight of 12 HEX-treated animals remained alive (resulting in an indeterminable median survival time) while one of 12 vehicle-treated rodents remained, yielding a median survival time of 10.9 days. Brains of six of the eight survivors were positive for amoebae, suggesting the agent at the tested dose suppressed, but did not eliminate, infection. These findings suggest that HEX is a promising lead for the treatment of PAM.

biochemistry↗

bioRxiv2022

Regulation of multiple dimeric states of E-cadherin by adhesion activating antibodies revealed through Cryo-EM and X-ray crystallography

E-cadherin adhesion is regulated at the cell surface, a process that can be replicated by activating antibodies. We use cryo-EM and X-ray crystallography to examine functional states of the cadherin adhesive dimer. This dimer is mediated by N-terminal beta strand-swapping involving Trp2, and forms via a different transient X-dimer intermediate. X-dimers are observed in cryo-EM along with monomers and strand-swap dimers, indicating that X-dimers form stable interactions. A novel EC4-mediated dimer was also observed. Activating Fab binding caused no gross structural changes in E- cadherin monomers but can facilitate strand swapping. Moreover, activating Fab binding is incompatible with the formation of the X-dimer. Both cryo-EM and X-ray crystallography reveal a distinctive twisted strand-swap dimer conformation caused by an outward shift in the N-terminal beta strand that may represent a strengthened state. Thus, regulation of adhesion involves changes in cadherin dimer configurations.

biochemistry↗

bioRxiv2021

Structures of plasmepsin X from P. falciparum reveal a novel inactivation mechanism of the zymogen and molecular basis for binding of inhibitors in mature enzyme

Plasmodium falciparum plasmepsin X (PfPMX), involved in the invasion and egress of this deadliest malarial parasite, is essential for its survival and hence considered as an important drug target. We report the first crystal structure of PfPMX zymogen containing a novel fold of its prosegment. A unique twisted loop from the prosegment and arginine 244 from the mature enzyme are involved in zymogen inactivation; such mechanism, not previously reported, might be common for apicomplexan proteases similar to PfPMX. The maturation of PfPMX zymogen occurs through cleavage of its prosegment at multiple sites. Our data provide thorough insights into the mode of binding of a substrate and a potent inhibitor 49c to PfPMX. We present molecular details of inactivation, maturation, and inhibition of PfPMX that should aid in the development of potent inhibitors against pepsin-like aspartic proteases from apicomplexan parasites.

biophysics↗

bioRxiv2020

Naegleria fowleri: protein structures to facilitate drug discovery for the deadly, pathogenic free-living amoeba

Naegleria fowleri is a pathogenic, thermophilic, free-living amoeba which causes primary amebic meningoencephalitis (PAM). Penetrating the olfactory mucosa, the brain-eating amoeba travels along the olfactory nerves, burrowing through the cribriform plate to its destination: the brains frontal lobes. The amoeba thrives in warm, freshwater environments, with peak infection rates in the summer months and has a mortality rate of approximately 97%. A major contributor to the pathogens high mortality is the lack of sensitivity of N. fowleri to current drug therapies, even in the face of combination-drug therapy. To enable rational drug discovery and design efforts we have pursued protein production and crystallography-based structure determination efforts for likely drug targets from N. fowleri. N. fowleri genes were selected if they had homology to drug targets listed in Drug Bank or were nominated by primary investigators engaged in N. fowleri research. In 2017, 178 N. fowleri protein targets were queued to the Seattle Structural Genomics Center of Infectious Disease (SSGCID) pipeline, and to date 89 soluble recombinant proteins and 19 unique target structures have been produced. Many of the new protein structures are potential drug targets and contain structural differences compared to their human homologs, which could allow for the development of pathogen-specific inhibitors. Five of the structures were analyzed in more detail, and four of five show promise that selective inhibitors of the active site could be found. The 19 solved crystal structures build a foundation for future work in combating this devastating disease by encouraging further investigation to stimulate drug discovery for this neglected pathogen.

molecular biology↗

bioRxiv2018

Enzymatic and structural characterization of the Naegleria fowleri glucokinase

Infection with the free-living amoeba Naegleria fowleri leads to life-threatening primary amoebic meningoencephalitis. Efficacious treatment options for these infections are limited and the mortality rate is very high (~98%). Parasite metabolism may provide suitable targets for therapeutic design. Like most other organisms, glucose metabolism is critical for parasite viability, being required for growth in culture. The genome of the parasite encodes a single glucose phosphorylating enzyme, a glucokinase (Glck). The products of this enzyme are required for both glycolysis and the pentose phosphate pathway. The N. fowleri Glck (NfGlck) shares limited (25%) amino acid identity with the mammalian host enzyme (HsGlck), suggesting that parasite-specific inhibitors with anti-amoeba activity could be generated. Following heterologous expression, NfGlck was found to have a limited hexose substrate range, with greatest activity observed with glucose. The enzyme had apparent Km values of 42.5 {+/-} 7.3 M and 141.6 {+/-} 9.9 M for glucose and ATP, respectively. The NfGlck structure was determined and refined to 2.2 [A] resolution, revealing that the enzyme shares greatest structural similarity with the Trypanosoma cruzi Glck. These similarities include binding modes and binding environments for substrates. To identify inhibitors of NfGlck, we screened a small collection of inhibitors of glucose phosphorylating enzymes and identified several small molecules with IC50 values < 1 M that may prove useful as hit chemotypes for further lead and therapeutic development against N. fowleri.

biochemistry↗