Pyrroloquinoline quinone (PQQ) is a peptide-derived redox cofactor produced by prokaryotes that also plays beneficial roles in organisms from other kingdoms. We review recent developments on the pathway of PQQ biogenesis, focusing on the mechanisms of PqqE, PqqF/G, and PqqB. These advances may shed light on other, uncharacterized biosynthetic pathways.

The lithium-sulfur cell has shown great prospects for future energy conversion and storage systems due to the high theoretical specific capacity of sulfur, 1675 mAh g−1. However, it has been hindered by rapid capacity decay and low energy efficiency. In this work, polypyrrole (PPy)/TiO2 nanotubes with coaxial heterogeneous structure as the substrate of the cathode is prepared and used to improve the electrochemical performance of sulfur electrodes. TiO2 nanotubes decorated with PPy provide a highly ordered conductive framework for Li+ ion diffusion and reaction with sulfur. This architecture also is helpful for trapping the produced polysulfides, and as a result attenuates the capacity decay. Furthermore, the heat treatment temperature used in the sulfur loading process has been confirmed to have an important impact on the overall performance of the resultant cell. The as-designed S/PPy/TiO2 nanotube cathode using an elevated heating temperature shows excellent cycling stability with a high discharge capacity of 1150 mAh g−1 and average coulombic efficiency of 96% after 100 cycles.

Oxalate decarboxylase (OxDC) catalyzes the disproportionation of oxalic acid monoanion into CO2 and formate. The enzyme has long been hypothesized to utilize dioxygen to form mononuclear Mn(III) or Mn(IV) in the catalytic site during turnover. Recombinant OxDC, however, contains only tightly bound Mn(II), and direct spectroscopic detection of the metal in higher oxidation states under optimal catalytic conditions (pH 4.2) has not yet been reported. Using parallel mode electron paramagnetic resonance spectroscopy, we now show that substantial amounts of Mn(III) are indeed formed in OxDC, but only in the presence of oxalate and dioxygen under acidic conditions. These observations provide the first direct support for proposals in which Mn(III) removes an electron from the substrate to yield a radical intermediate in which the barrier to C-C bond cleavage is significantly decreased. Thus, OxDC joins a small list of enzymes capable of stabilizing and controlling the reactivity of the powerful oxidizing species Mn(III).

Pyrroloquinoline quinone (PQQ) is an important redox active quinocofactor produced by a wide variety of bacteria. A key step in PQQ biosynthesis is a carbon-carbon cross-link reaction between glutamate and tyrosine side chains within the ribosomally synthesized peptide substrate PqqA. This reaction is catalyzed by the radical SAM enzyme PqqE. Previous X-ray crystallographic and spectroscopic studies suggested that PqqE, like the other members of the SPASM domain family, contains two auxiliary Fe-S clusters (AuxI and AuxII) in addition to the radical SAM [4Fe-4S] cluster. However, a clear assignment of the electron paramagnetic resonance (EPR) signal of each Fe-S cluster was hindered by the isolation of a His₆-tagged PqqE variant with an altered AuxI cluster. In this work, we are able to isolate soluble PqqE variants by using a less disruptive strep-tactin chromatographic approach. We have unambiguously identified the EPR signatures for four forms of Fe-S clusters present in PqqE through the use of multifrequency EPR spectroscopy: the RS [4Fe-4S] cluster, the AuxII [4Fe-4S] cluster, and two different clusters ([4Fe-4S] and [2Fe-2S]) bound in the AuxI site. The RS [4Fe-4S] cluster, the AuxII [4Fe-4S] cluster, and the [2Fe-2S] cluster form in the AuxI site can all be reduced by sodium dithionite, with g tensors of their reduced form determined as [2.040, 1.927, 1.897], [2.059, 1.940, 1.903], and [2.004, 1.958, 1.904], respectively. The AuxI [4Fe-4S] cluster that is determined on the basis of its relaxation profile can be reduced only by using low-potential reductants such as Ti(III) citrate or Eu(II)-DTPA to give rise to a g₁ = 2.104 signal. Identification of the EPR signature for each cluster paves the way for further investigations of SPASM domain radical SAM enzymes.

Interactions among proteins and peptides are essential for many biological activities including the tailoring of peptide substrates to produce natural products. The first step in the production of the bacterial redox cofactor pyrroloquinoline quinone (PQQ) from its peptide precursor is catalyzed by a radical SAM (rSAM) enzyme, PqqE. We describe the use of hydrogen-deuterium exchange mass spectrometry (HDX-MS) to characterize the structure and conformational dynamics in the protein-protein and protein-peptide complexes necessary for PqqE function. HDX-MS-identified hotspots can be discerned in binary and ternary complex structures composed of the peptide PqqA, the peptide-binding chaperone PqqD, and PqqE. Structural conclusions are supported by size-exclusion chromatography coupled to small-angle X-ray scattering (SEC-SAXS). HDX-MS further identifies reciprocal changes upon the binding of substrate peptide and S-adenosylmethionine (SAM) to the PqqE/PqqD complex: long-range conformational alterations have been detected upon the formation of a quaternary complex composed of PqqA/PqqD/PqqE and SAM, spanning nearly 40 Å, from the PqqA binding site in PqqD to the PqqE active site Fe₄S₄. Interactions among the various regions are concluded to arise from both direct contact and distal communication. The described experimental approach can be readily applied to the investigation of protein conformational communication among a large family of peptide-modifying rSAM enzymes.

PqqE is the first enzyme in the biosynthetic pathway of the redox cofactor pyrroloquinoline quinone (PQQ), catalyzing the formation of a carbon-carbon bond in the precursor peptide PqqA. PqqE is a radical S-adenosyl-l-methionine (SAM) (RS) enzyme, a family of enzymes that use the reductive cleavage of a [4Fe-4S] cluster-bound SAM molecule to generate a 5'-deoxyadenosyl radical. This radical is then used to initiate an array of reactions that otherwise would be unlikely to occur. PqqE is a founding member of a subset family of RS enzymes that, additionally to the SAM [4Fe-4S] cluster, have a SPASM domain containing additional, auxiliary Fe-S clusters. Most radical SAM enzymes are highly sensitive to oxygen, which destroys their Fe-S clusters. This can pose several limitations when working with these enzymes, since most of the work has to be done under anaerobic conditions. Here, we summarize the methods developed in our lab for the expression and purification of PqqE. We also highlight the several methods we have used for the characterization of the enzyme.

Quantitative susceptibility mapping (QSM) is increasingly used to measure variation in tissue composition both in the brain and in other areas of the body in a range of disease pathologies. Although QSM measurements were originally believed to be independent of the echo time (TE) used in the gradient-recalled echo (GRE) acquisition from which they are derived; recent literature (Sood et al., 2016) has shown that these measurements can be highly TE-dependent in a number of brain regions. In this work we systematically investigate possible causes of this effect through analysis of apparent frequency and QSM measurements derived from data acquired at multiple TEs in vivo in healthy brain regions and in cerebral microbleeds (CMBs); QSM data acquired in a gadolinium-doped phantom; and in QSM data derived from idealized simulated phase data. Apparent frequency measurements in the optic radiations (OR) and central corpus callosum (CC) were compared to those predicted by a 3-pool white matter model, however the model failed to fully explain contrasting frequency profiles measured in the OR and CC. Our results show that TE-dependent QSM measurements can be caused by a failure of phase unwrapping algorithms in and around strong susceptibility sources such as CMBs; however, in healthy brain regions this behavior appears to result from intrinsic non-linear phase evolution in the MR signal. From these results we conclude that care must be taken when deriving frequency and QSM measurements in strong susceptibility sources due to the inherent limitations in phase unwrapping; and that while signal compartmentalization due to tissue microstructure and content is a plausible cause of TE-dependent frequency and QSM measurements in healthy brain regions, better sampling of the MR signal and more complex models of tissue are needed to fully exploit this relationship.

This is the report of a 76-year-old male with typical lesions of acanthosis nigricans maligna (ANM), florid cutaneous papillomatosis (FCP), and tripe palms (TP) for 2 years. He did not have any gastrointestinal complaints. Pathologic findings of skin supported the diagnosis of ANM. Because gastric adenocarcinoma is the most common neoplasm associated with these paraneoplastic dermatoses, further tests were carried out. Endoscopic examination was performed and an adenocarcinoma of the esophagogastric junction was confirmed. Meanwhile, multiple small polyps in the middle and the lower thirds of the esophagus were observed. The patient was referred for further evaluation and subsequent surgical resection of the tumor.Acanthosis nigricans (AN) is a hyperkeratotic mucocutaneous eruption of heterogenous etiology, which is characterized by hyperpigmentation, velvety cutaneous thickening, intensified skin markings, and development of verrucous excrescences typically involving the intertriginous areas. AN is classified into benign and malignant forms on the basis of clinical associations. Malignant acanthosis nigricans (MAN) tends to be extensive and involves mucosal surfaces, mostly in elderly people. Florid cutaneous papillomatosis (FCP), also known as the Schwartz-Burgess syndrome, is characterized by the rapid appearance of multiple verrucous lesions that are clinically indistinguishable from common warts [1]. Tripe palms (TP) is characterized by diffuse, yellowish palmar hyperkeratosis, with enhancement of the epidermal ridges on the hands (dermatoglyphics), resembling intestinal villosities [1]. The association of these three paraneoplastic dermatoses (FCP, ANM and TP) in the same patient has been reported. Herein, we report an elderly male with three paraneoplastic dermatoses for two years. On the initial presentation, he did not report any systemic complaints; diagnostic tests confirmed the presence of a gastric adenocarcinoma.

The human brain rapidly develops during the first two years following birth. Quantitative susceptibility mapping (QSM) provides information of iron and myelin variations. It is considered to be a valuable tool for studying brain development in early life. In the present work, QSM is performed on neonates, 1-year and 2-year old infants, as well as a group of adults for the purpose of reference. Age-specific templates representing common brain structures are built for each age group. The neonate and infant QSM templates have shown some unique findings compared to conventional T1w and T2w imaging techniques. The contrast between the gray and white matters on the QSM images did not change through brain development from neonate to adult. A linear correlation was found between brain myelination determined in this study and the microscopic myelin degree determined by a previous autopsy study. Also, the magnetic susceptibility values of the cerebral spinal fluid (CSF) exhibit a gradually decreasing trend from birth to 2 years old and to adulthood. The findings suggest that the macromolecular content, myelin, and iron may play the most important contributing factors for the magnetic susceptibility of neonate and infant brain. QSM can be a powerful means to study early brain development and related pathologies that involve alterations in macromolecular content, iron, or brain myelination.

Understanding the relationship between the metallocofactor and its protein environment is the key to uncovering the mechanism of metalloenzymes. PqqE, a radical S-adenosylmethionine enzyme in pyrroloquinoline quinone (PQQ) biosynthesis, contains three iron-sulfur cluster binding sites. Two auxiliary iron-sulfur cluster binding sites, designated as AuxI and AuxII, use distinctive ligands compared to other proteins in the family while their functions remain unclear. Here, we investigate the electronic properties of these iron-sulfur clusters and compare the catalytic efficiency of wild-type (WT) Methylorubrum extorquens AM1 PqqE to a range of mutated constructs. Using native mass spectrometry, protein film electrochemistry, and electron paramagnetic resonance spectroscopy, we confirm the previously proposed incorporation of a mixture of [2Fe-2S] and [4Fe-4S] clusters at the AuxI site and are able to assign redox potentials to each of the three iron-sulfur clusters. Significantly, a conservative mutation at AuxI, C268H, shown to selectively incorporate a [4Fe-4S] cluster, catalyzes an enhancement of uncoupled S-adenosylmethionine cleavage relative to WT, together with the elimination of detectable peptide cross-linked product. While a [4Fe-4S] cluster can be tolerated at the AuxI site, the aggregate findings suggest a functional [2Fe-2S] configuration within the AuxI site. PqqE variants with nondestructive ligand replacements at AuxII also show that the reduction potential at this site can be manipulated by changing the electronegativity of the unique aspartate ligand. A number of novel mechanistic features are proposed based on the kinetic and spectroscopic data. Additionally, bioinformatic analyses suggest that the unique ligand environment of PqqE may be relevant to its role in PQQ biosynthesis within an oxygen-dependent biosynthetic pathway.