Structural Changes during the Photorepair and Binding Processes of (6-4) Photolyase with (6-4) Photoproducts in Single- and Double-Stranded DNA.

Photolyases (PHRs) repair ultraviolet (UV)-induced DNA photoproducts into normal bases. In this study, we measured the conformational changes upon photoactivation and photorepair processes of a PHR and its specific substrates, (6-4)PHR and a pyrimidine(6-4)pyrimidone photoproduct ((6-4)PP), by light-induced difference Fourier transform infrared (FT-IR) spectroscopy. The single-stranded DNA with (6-4)PP (ss(6-4)PP) was used as a substrate and the resultant FT-IR spectra were compared with the previous results on double-stranded DNA with (6-4)PP (ds(6-4)PP).

Addressing Structural Racism Is Critical for Ameliorating the Childhood Obesity Epidemic in Black Youth.

Racism and childhood obesity are both pervasive factors adversely affecting the health and wellbeing of children and adolescents in the United States. The association between racism and obesity has been touched upon in the literature; yet most work has focused on a few dimensions of intersectionality of these two domains at one time. The renewed focus on structural racism as the primary contributor to distress of Black individuals in the United States has highlighted the urgency of identifying the contributions of racism to the childhood obesity epidemic.

ATP binding promotes light-induced structural changes to the protein moiety of cryptochrome 1.

Cryptochromes (CRYs) are blue-light receptors involved in photomorphogenesis in plants. Flavin adenine dinucleotide (FAD) is one of the chromophores of cryptochromes; its resting state oxidized form is converted into a signalling state neutral semiquionod radical (FADH˙) form. Studies have shown that cryptochrome 1 from Arabidopsis thaliana (AtCRY1) can bind ATP at its photolyase homology region (PHR), resulting in accumulation of FADH˙ form.

A Self-Assisting Protein Folding Model for Teaching Structural Molecular Biology.

Structural molecular biology is now becoming part of high school science curriculum thus posing a challenge for teachers who need to convey three-dimensional (3D) structures with conventional text and pictures. In many cases even interactive computer graphics does not go far enough to address these challenges. We have developed a flexible model of the polypeptide backbone using 3D printing technology.

Pediatric Weight Management Program Outcomes in a Largely Minority, Low Socioeconomic Status Population.

This article describes the outcomes of a pediatric weight management program for a population primarily composed of minority ethnic groups and those from a lower socioeconomic status group. As these groups are disproportionally affected by pediatric obesity and overweight complicated by higher rates of attrition and poorer response to intervention, it is important that adequate and effective treatment exists for patients in these groups. Further research is needed to analyze the outcomes and attrition in these high-risk populations.

Self-Assembly of the Cephalopod Protein Reflectin.

Films from the cephalopod protein reflectin demonstrate multifaceted functionality as infrared camouflage coatings, proton transport media, and substrates for growth of neural stem cells. A detailed study of the in vitro formation, structural characteristics, and stimulus response of such films is presented. The reported observations hold implications for the design and development of advanced cephalopod-inspired functional materials.

Functional Conversion of CPD and (6-4) Photolyases by Mutation.

Ultraviolet (UV) light from the sun damages DNA by forming a cyclobutane pyrimidine dimer (CPD) and pyrimidine(6-4)pyrimidone photoproducts [(6-4) PP]. Photolyase (PHR) enzymes utilize near-UV/blue light for DNA repair, which is initiated by light-induced electron transfer from the fully reduced flavin adenine dinucleotide chromophore. Despite similar structures and repair mechanisms, the functions of PHR are highly selective; CPD PHR repairs CPD, but not (6-4) PP, and vice versa.

Single Hydrogen Bond Donation from Flavin N5 to Proximal Asparagine Ensures FAD Reduction in DNA Photolyase.

The spread of the absorbance of the stable FADH(•) radical (300-700 nm) allows CPD photolyase to highly efficiently form FADH(-), making it functional for DNA repair. In this study, FTIR spectroscopy detected a strong hydrogen bond, from FAD N5-H to the carbonyl group of the Asn378 side chain, that is modulated by the redox state of FAD. The observed characteristic frequency shifts were reproduced in quantum-mechanical models of the flavin binding site, which were then employed to elucidate redox tuning governed by Asn378.

Structural Changes of the Active Center during the Photoactivation of Xenopus (6-4) Photolyase.

Photolyases (PHRs) repair the UV-induced photoproducts, cyclobutane pyrimidine dimer (CPD) or pyrimidine-pyrimidone (6-4) photoproduct [(6-4) PP], restoring normal bases to maintain genetic integrity. CPD and (6-4) PP are repaired by substrate-specific PHRs, CPD PHR and (6-4) PHR, respectively. Flavin adenine dinucleotide (FAD) is the chromophore of both PHRs, and the resting oxidized form (FAD(ox)), at least under in vitro purified conditions, is first photoconverted to the neutral semiquinoid radical (FADH(•)) form, followed by photoconversion into the enzymatically active fully reduced (FADH(-)) form.

Structural, Functional, and Immunogenic Insights on Cu,Zn Superoxide Dismutase Pathogenic Virulence Factors from Neisseria meningitidis and Brucella abortus.

Unlabelled: Bacterial pathogens Neisseria meningitidis and Brucella abortus pose threats to human and animal health worldwide, causing meningococcal disease and brucellosis, respectively. Mortality from acute N. meningitidis infections remains high despite antibiotics, and brucellosis presents alimentary and health consequences.

Reaction dynamics of the UV-B photosensor UVR8.

UVR8 is a recently discovered ultraviolet-B (UV-B) photoreceptor protein identified in plants and algae. In the dark state, UVR8 exists as a homodimer, whereas UV-B irradiation induces UVR8 monomerization and initiation of signaling. Although the biological functions of UVR8 have been studied, the fundamental reaction mechanism and associated kinetics have not yet been fully elucidated.

Structural role of two histidines in the (6-4) photolyase reaction.

Photolyases (PHRs) are DNA repair enzymes that revert UV-induced photoproducts, either cyclobutane pyrimidine dimers (CPD) or (6-4) photoproducts (PPs), into normal bases to maintain genetic integrity. (6-4) PHR must catalyze not only covalent bond cleavage, but also hydroxyl or amino group transfer, yielding a more complex mechanism than that postulated for CPD PHR. Previous mutation analysis revealed the importance of two histidines in the active center, H354 and H358 for Xenopus (6-4) PHR, whose mutations significantly lowered the enzymatic activity.

FTIR study of CPD photolyase with substrate in single strand DNA.

Photolyases (PHRs) utilize near UV/blue light to specifically repair the major photoproducts (PPs) of UV-induced damaged DNA. The cyclobutane pyrimidine dimer (CPD)-PHR binds flavin adenine dinucleotide (FAD) as a cofactor and repairs CPD lesions in double-stranded DNA. To understand the activation and repair mechanism of CPD-PHR, we applied light-induced difference Fourier transform infrared (FTIR) spectroscopy to CPD-PHR, whose signals were identified by use of isotope-labeling.

Aggregation propensities of superoxide dismutase G93 hotspot mutants mirror ALS clinical phenotypes.

Protein framework alterations in heritable Cu, Zn superoxide dismutase (SOD) mutants cause misassembly and aggregation in cells affected by the motor neuron disease ALS. However, the mechanistic relationship between superoxide dismutase 1 (SOD1) mutations and human disease is controversial, with many hypotheses postulated for the propensity of specific SOD mutants to cause ALS. Here, we experimentally identify distinguishing attributes of ALS mutant SOD proteins that correlate with clinical severity by applying solution biophysical techniques to six ALS mutants at human SOD hotspot glycine 93.

Copper-based pulsed dipolar ESR spectroscopy as a probe of protein conformation linked to disease states.

We demonstrate the ability of pulsed dipolar electron spin resonance (ESR) spectroscopy (PDS) to report on the conformation of Cu-Zn superoxide dismutase (SOD1) through the sensitive measurement of dipolar interactions between inherent Cu(2+) ions. Although the extent and the anisotropy of the Cu ESR spectrum provides challenges for PDS, Ku-band (17.3 GHz) double electron-electron resonance and double-quantum coherence variants of PDS coupled with distance reconstruction methods recover Cu-Cu distances in good agreement with crystal structures.

Flavin adenine dinucleotide chromophore charge controls the conformation of cyclobutane pyrimidine dimer photolyase α-helices.

Observations of light-receptive enzyme complexes are usually complicated by simultaneous overlapping signals from the chromophore, apoprotein, and substrate, so that only the initial, ultrafast, photon-chromophore reaction and the final, slow, protein conformational change provide separate, nonoverlapping signals. Each provides its own advantages, whereas sometimes the overlapping signals from the intervening time scales still cannot be fully deconvoluted. We overcome the problem by using a novel method to selectively isotope-label the apoprotein but not the flavin adenine dinucleotide (FAD) cofactor.

ATP binding turns plant cryptochrome into an efficient natural photoswitch.

Cryptochromes are flavoproteins that drive diverse developmental light-responses in plants and participate in the circadian clock in animals. Plant cryptochromes have found application as photoswitches in optogenetics. We have studied effects of pH and ATP on the functionally relevant photoreduction of the oxidized FAD cofactor to the semi-reduced FADH(·) radical in isolated Arabidopsis cryptochrome 1 by transient absorption spectroscopy on nanosecond to millisecond timescales.

FRET-based reporters for the direct visualization of abscisic acid concentration changes and distribution in Arabidopsis.

Abscisic acid (ABA) is a plant hormone that regulates plant growth and development and mediates abiotic stress responses. Direct cellular monitoring of dynamic ABA concentration changes in response to environmental cues is essential for understanding ABA action. We have developed ABAleons: ABA-specific optogenetic reporters that instantaneously convert the phytohormone-triggered interaction of ABA receptors with PP2C-type phosphatases to send a fluorescence resonance energy transfer (FRET) signal in response to ABA.

Variable electron transfer pathways in an amphibian cryptochrome: tryptophan versus tyrosine-based radical pairs.

Electron transfer reactions play vital roles in many biological processes. Very often the transfer of charge(s) proceeds stepwise over large distances involving several amino acid residues. By using time-resolved electron paramagnetic resonance and optical spectroscopy, we have studied the mechanism of light-induced reduction of the FAD cofactor of cryptochrome/photolyase family proteins.

Detection of distinct α-helical rearrangements of cyclobutane pyrimidine dimer photolyase upon substrate binding by Fourier transform infrared spectroscopy.

Photolyases (PHRs) utilize near-ultraviolet (UV)-blue light to specifically repair the major photoproducts (PPs) of UV-induced damaged DNA. The cyclobutane pyrimidine dimer PHR (CPD-PHR) from Escherichia coli binds flavin adenine dinucleotide (FAD) as a cofactor and 5,10-methenyltetrahydrofolate as a light-harvesting pigment and specifically repairs CPD lesions. By comparison, a second photolyase known as (6-4) PHR, present in a range of higher organisms, uniquely repairs (6-4) PPs.

Amyotrophic lateral sclerosis: update and new developments.

Amyotrophic lateral sclerosis (ALS) is the most common form of motor neuron disease. It is typically characterized by adult-onset degeneration of the upper and lower motor neurons, and is usually fatal within a few years of onset. A subset of ALS patients has an inherited form of the disease, and a few of the known mutant genes identified in familial cases have also been found in sporadic forms of ALS.

Fourier-transform infrared study of the photoactivation process of Xenopus (6-4) photolyase.

Photolyases (PHRs) are blue light-activated DNA repair enzymes that maintain genetic integrity by reverting UV-induced photoproducts into normal bases. The flavin adenine dinucleotide (FAD) chromophore of PHRs has four different redox states: oxidized (FAD(ox)), anion radical (FAD(•-)), neutral radical (FADH(•)), and fully reduced (FADH(-)). We combined difference Fourier-transform infrared (FTIR) spectroscopy with UV-visible spectroscopy to study the detailed photoactivation process of Xenopus (6-4) PHR.

Structural tuning of the fluorescent protein iLOV for improved photostability.

Fluorescent proteins derived from light, oxygen, or voltage (LOV) domains offer advantages over green fluorescent protein (GFP) from their small size and efficacy under anaerobic conditions. The flavoprotein improved LOV (iLOV) was engineered from the blue light receptor phototropin as a reporter of viral infection. To inform the molecular basis for the improved, photoreversible, fluorescent properties of iLOV, we employed directed evolution and determined five LOV crystallographic structures.

Plant UVR8 photoreceptor senses UV-B by tryptophan-mediated disruption of cross-dimer salt bridges.

The recently identified plant photoreceptor UVR8 (UV RESISTANCE LOCUS 8) triggers regulatory changes in gene expression in response to ultraviolet-B (UV-B) light through an unknown mechanism. Here, crystallographic and solution structures of the UVR8 homodimer, together with mutagenesis and far-UV circular dichroism spectroscopy, reveal its mechanisms for UV-B perception and signal transduction. β-propeller subunits form a remarkable, tryptophan-dominated, dimer interface stitched together by a complex salt-bridge network.