Lateral flow assays: Progress and evolution of recent trends in point-of-care applications.

Paper based point-of-care (PoC) detection platforms applying lateral flow assays (LFAs) have gained paramount approval in the diagnostic domain as well as in environmental applications owing to their ease of utility, low cost, and rapid signal readout. It has centralized the aspect of self-evaluation exhibiting promising potential in the last global pandemic era of Covid-19 implementing rapid management of public health in remote areas. In this perspective, the present review is focused towards landscaping the current framework of LFAs along with integration of components and characteristics for improving the assay by pushing the detection limits. The review highlights the synergistic aspects of assay designing, sample enrichment strategies, novel nanomaterials-based signal transducers, and high-end analytical techniques that contribute significantly towards sensitivity and specificity enhancement. Various recent studies are discussed supporting the innovations in LFA systems that focus upon the accuracy and reliability of rapid PoC testing. The review also provides a comprehensive overview of all the possible difficulties in commercialization of LFAs subjecting its applicability to pathogen surveillance, water and food testing, disease diagnostics, as well as to agriculture and environmental issues.

Bromodomain protein BRD4 directs mitotic cell division of mouse fibroblasts by inhibiting DNA damage.

Bromodomain protein BRD4 binds to acetylated histones to regulate transcription. BRD4 also drives cancer cell proliferation. However, the role of BRD4 in normal cell growth has remained unclear. Here, we investigated this question by using mouse embryonic fibroblasts with conditional Brd4 knockout (KO). We found that Brd4KO cells grow more slowly than wild type cells; they do not complete replication, fail to achieve mitosis, and exhibit extensive DNA damage throughout all cell cycle stages. BRD4 was required for expression of more than 450 cell cycle genes including genes encoding core histones and centromere/kinetochore proteins that are critical for genome replication and chromosomal segregation. Moreover, we show that many genes controlling R-loop formation and DNA damage response (DDR) require BRD4 for expression. Finally, BRD4 constitutively occupied genes controlling R-loop, DDR and cell cycle progression. In summary, BRD4 epigenetically marks above genes and serves as a master regulator of normal cell growth.

FibroScan as a surveillance marker post-cavopulmonary anastomosis for single-ventricle physiology.

Purpose: Single-ventricle physiology encompasses a group of congenital cardiac malformations with only one functional ventricle. The Fontan procedure is the final palliation of this pathway and has its complications. One of these is Fontan-associated liver disease (FALD). It is known that all patients with Fontan will have FALD, due to the physiology of the Fontan circuit, and only the severity will vary. The pathophysiology of hepatic damage in FALD is unique and not easily detectable by routine non-invasive investigations. Therefore, this study is aimed to identify if FibroScan can be used as a surveillance marker to detect and assess the progression of FALD. Methods: Patients who attended the Cardiothoracic and Vascular Surgery Outpatient Department (OPD) for follow-up post-cavopulmonary anastomosis (bidirectional Glenn and Fontan) were enrolled in this study. They underwent routine examinations and tests, and in addition a FibroScan was performed. Results: FibroScan showed that the liver stiffness measurement (LSM) was increased in all patients who had undergone Fontan and a couple of patients who had undergone bidirectional (BD) Glenn. The LSM was 12.29 (± 3.59) kPa in patients post-Fontan and 6.64 (± 4.24) kPa in patients post-BD Glenn. This raised LSM was not associated with a parallel rise in liver enzymes or other laboratory markers associated with liver function. This emphasizes the suitability of FibroScan as an early and non-invasive surveillance tool for monitoring the progression of hepatic venous congestion and FALD. Conclusion: We have found that LSM via FibroScan can effectively be a surveillance or screening test for FALD. Serial FibroScans can be used to monitor the progress of liver disease. Raised LSM may also have a role in predicting the morbidity for completion of Fontan post-BD Glenn operation. Large-scale studies are needed to validate our findings.

Gold Nanoparticle-based Biosensors for Point-of-Care Diagnostics: A Review of Sensing Nanoparticle Applications and Future Prospects.

Point of Care Diagnostics (POCD) is quintessential in hospitals and the healthcare sector as the secants uplift the quality of medical care and the life of a patient by facilitating quick identification of the underlying pathological condition. Nanotechnology can provide opportunities and has potential in the development of new-age sensing/diagnostic tools. Owing to extraordinary features (e.g., higher density, effective catalysis, good conduction, biocompatibility, inertness, and greater surface-to-volume ratio), gold nanoparticles (AuNPs) are frequently employed in POCT (Point-of-Care-Testing). Gold nanoparticles-based colorimetric methods are widely used in the rapid, sensitive, and selective detection of analytes/target molecules. AuNPs description is critical for their possible utility in prophylaxis, diagnostics, and treatment of an ailment. AuNPs interact with organic/inorganic target molecules to generate colorimetric shift that enables the accurate, precise, and subtle recognition of biologicals (e.g., microorganisms, cellular components, and proteins) and metal ions. This review focused on the need for AuNPs-based colorimetric application in prophylaxis, diagnostics, and treatment in healthcare and reviewed the future outlook of these AuNPs for biological applications. Different synthesis methods of AuNPs, their morphology, and characterization, including their surface functionalization, will be discussed in detail. AuNPs are very much preferable nanomaterials owing to exclusive optical, electrical, and photothermal features. AuNPs-based colorimetric biosensors are simple and possess great utility, yet these offer a robust technique to enable visual, quantitative analysis.

The Impact of microRNA SNPs on Breast Cancer: Potential Biomarkers for Disease Detection.

Breast cancer is considered a significant health concern worldwide, with genetic predisposition playing a critical role in its etiology. Single nucleotide polymorphisms (SNPs), particularly those within the 3' untranslated regions (3'UTRs) of target genes, are emerging as key factors in breast cancer susceptibility. Specifically, miRNAs have been recognized as possible novel approach for biomarkers discovery for both prognosis and diagnosis due to their direct association with cancer progression. Regional disparities in breast cancer incidence underscore the need for precise interventions, considering socio-cultural and economic factors. This review explores into the differential effects of SNP-miRNA interactions on breast cancer risk, emphasizing both risk-enhancing and protective associations across diverse populations. Furthermore, it explores the clinical implications of these findings, highlighting the potential of personalized approaches in breast cancer management. Additionally, it reviews the evolving therapeutic prospect of microRNAs (miRNAs), extending beyond cancer therapeutics to encompass various diseases, indicative of their versatility as therapeutic agents.

BRD4 directs mitotic cell division by inhibiting DNA damage.

BRD4 binds to acetylated histones to regulate transcription and drive cancer cell proliferation. However, the role of BRD4 in normal cell growth remains to be elucidated. Here we investigated the question by using mouse embryonic fibroblasts with conditional Brd4 knockout (KO). We found that Brd4KO cells grow more slowly than wild type cells: they do not complete replication, fail to achieve mitosis, and exhibit extensive DNA damage throughout all cell cycle stages. BRD4 was required for expression of more than 450 cell cycle genes including genes encoding core histones and centromere/kinetochore proteins that are critical for genome replication and chromosomal segregation. Moreover, we show that many genes controlling R-loop formation and DNA damage response (DDR) require BRD4 for expression. Finally, BRD4 constitutively occupied genes controlling R-loop, DDR and cell cycle progression. We suggest that BRD4 epigenetically marks those genes and serves as a master regulator of normal cell growth.

Conformational switching of aptamer biointerfacing graphene-gold nanohybrid for ultrasensitive label-free sensing of cardiac Troponin I.

The development of hybrid biofunctionalized nanomaterials has emerged as an attractive substitute for development of advanced biosensing platforms with superior synergistic properties. Herein, we report a label-free ultrasensitive electrochemical aptasensor comprising nanohybrid of graphene oxide (GO) and aptamer conjugated gold nanoparticles (GNP-A) for detection of cardiac biomarker Troponin I (TnI). The GNP-A are homogenously arranged by self-assembly on GO sheet to construct nanohybrid (GO@GNP-A) onto which the biomarker protein is analysed. TnI interactions at the aptamer biointerfaced nanohybrid surface causes electrochemical signal enhancement probed by using a redox active molecule. The consecutive increase in current signal is strongly attributed to conformational switching of aptamer and charge neutralization at the interface induced by TnI binding. The sensitivity of the nanohybrid aptasensor platform was found to be 0.001 pg/mL. The study has been further substantiated in Acute Myocardial Infarction (AMI) clinical samples for usage towards early, sensitive and efficient point-of-care detection of TnI.

Plant-mediated synthesis of silver nanoparticles: unlocking their pharmacological potential-a comprehensive review.

In recent times, nanoparticles have experienced a significant upsurge in popularity, primarily owing to their minute size and their remarkable ability to modify physical, chemical, and biological properties. This burgeoning interest can be attributed to the expanding array of biomedical applications where nanoparticles find utility. These nanoparticles, typically ranging in size from 10 to 100 nm, exhibit diverse shapes, such as spherical, discoidal, and cylindrical configurations. These variations are not solely influenced by the manufacturing processes but are also intricately linked to interactions with surrounding stabilizing agents and initiators. Nanoparticles can be synthesized through physical or chemical methods, yet the biological approach emerges as the most sustainable and eco-friendly alternative among the three. Among the various nanoparticle types, silver nanoparticles have emerged as the most encountered and widely utilized due to their exceptional properties. What makes the synthesis of silver nanoparticles even more appealing is the application of plant-derived sources as reducing agents. This approach not only proves to be cost-effective but also significantly reduces the synthesis time. Notably, silver nanoparticles produced through plant-mediated processes have garnered considerable attention in recent years due to their notable medicinal capabilities. This comprehensive review primarily delves into the diverse medicinal attributes of silver nanoparticles synthesized using plant-mediated techniques. Encompassing antimicrobial properties, cytotoxicity, wound healing, larvicidal effects, anti-angiogenesis activity, antioxidant potential, and antiplasmodial activity, the paper extensively covers these multifaceted roles. Additionally, an endeavor is made to provide an elucidated summary of the operational mechanisms underlying the pharmacological actions of silver nanoparticles.

Site-directed dual bioprobes inducing single-step nano-sandwich assay for the detection of cardiac troponin I.

Bioreceptor functionalized metallic nano-colloids have been identified as effective nanobioprobes to realize the detection of an analyte based on a common phenomenon of salt-induced aggregation. In marked contrast to this, we describe a nano-sandwich assay integrating the novel match-pair of aptamer and peptide functionalized gold nanoparticles. The site-directed biomolecular interaction of high affinity aptamer and peptide bioreceptors directed towards distinct sites of cardiac biomarker troponin I; this was found to form a nano-sandwich assay in a peculiar manner. The gold nanoconjugates interact with specific and distant regions of troponin I to result in collision of probes upon target identification. In the presence of TnI, both nanobioprobes bind at their respective sites forming a nano-sandwich pair providing a visual color change from red to blue. Thus, the presence of target TnI itself causes instant agglomeration in just a single-step without addition of any external aggregator. The assay imparts 100% specificity and 90% sensitivity in a dynamic concentration range of 0.1-500 ng/mL troponin I with detection limit as low as 0.084 ng/mL. The applicability of the assay has been validated in clinical samples of acute myocardial infarction patients thus establishing a promising point-of-care detection of TnI.

Hyperlipidemia impairs uterine ß-adrenergic signaling by reducing cAMP in late pregnant rats.

The aim of the present study was to reveal the effect of hyperlipidemia on ß2- and ß3-adrenergic signaling in late pregnant rat uterus. Hyperlipidemia was induced in female Wistar rats by feeding a high-fat high-cholesterol diet for 8 weeks before and after mating upto the 21st day of gestation. The effect of hyperlipidemia on ß-adrenergic signaling was studied with the help of tension experiments, real-time PCR and cAMP ELISA in 21-day pregnant rat uterus. In tension experiments, hyperlipidemia neither altered the spontaneous contractility nor the oxytocin-induced contractions. However, it decreased the -logEC50 values of ß2-adrenoceptor agonist, salbutamol and ß3-adrenoceptor agonist, BRL37344. It also decreased the efficacy of adenylyl cyclase activator, forskolin. Further, there was a significant decrease in salbutamol and BRL37344-stimulated cAMP content in uterine tissues. However, there was no alteration in mRNA expressions of ß2-adrenoceptor (Adrb2), ß3-adrenoceptor (Adrb3) and Gs protein (Gnas) though there was a significant increase in the mRNA expression of Gi protein (Gnai). In conclusion, reduced cAMP content after beta-adrenergic receptor stimulation, which correlates with an increase in Gnai mRNA, may explain the mechanism of the impairment of uterine ß-adrenergic signaling in hyperlipidemic pregnant rats. The clinical implication of the present study may relate to reduced myometrial relaxant response to ß-adrenergic agonists in high fat-induced uterine dysfunction.

Reverse genetic analysis of yeast YPR099C/MRPL51 reveals a critical role of both overlapping ORFs in respiratory growth and MRPL51 in mitochondrial DNA maintenance.

The Saccharomyces cerevisiae genome contains 6572 ORFs, of which 680 ORFs are classified as dubious ORFs. A dubious ORF is a small, noncoding, nonconserved ORF that overlaps with another ORF of the complementary strand. Our study characterizes a dubious/nondubious ORF pair, YPR099C/MRPL51, and shows the transcript and protein level expression of YPR099C. Its subcellular localization was observed in the mitochondria. The overlapping ORF, MRPL51, encodes a mitochondrial ribosomal protein of large subunit. Deletion of any ORF from YPR099C/MRPL51 pair induces common phenotypes, i.e. loss of mtDNA, lack of mitochondrial fusion and lack of respiratory growth, due to the double deletion (ypr099cΔ/Δmrpl51Δ/Δ) caused by sequence overlap. Hence, we created the single deletions of each ORF of the YPR099C/MRPL51 pair by an alternative approach to distinguish their phenotypes and identify the specific functions. Both the ORFs were found essential for the functional mitochondria and respiratory growth, but MRPL51 showed its specific requirement in mtDNA stability. The mechanism of mtDNA maintenance by Mrpl51 is probably Mhr1 dependent that physically interacts with Mrpl51 and also regulates mtDNA repair. Overall, our study provides strong evidence for the protein level expression of a dubious ORF YPR099C and the bifunctional role of Mrpl51 in mtDNA maintenance.

The Crg1 N-Terminus Is Essential for Methyltransferase Activity and Cantharidin Resistance in Saccharomyces cerevisiae.

Crg1 is an S-adenosylmethionine (SAM)-dependent methyltransferase required for cantharidin resistance in yeast. Crg1 has a well-characterized methyltransferase domain that inactivates cantharidin by methylation. However, the remaining part of the Crg1 protein is yet to be functionally characterized. In this study, we identified an essential role of the N-terminus of Crg1 in methyltransferase activity and cantharidin resistance. Yeast cells lacking 41 residues of the N-terminus of Crg1 ( crg1ΔN) showed hypersensitivity to cantharidin as same as the null mutant, crg1. The mass spectrometry-based biochemical enzyme assay revealed a loss of methyltransferase activity in Crg1ΔN, which justifies the loss of cantharidin resistance, as well. The subcellular distribution of Crg1ΔN-daGFP showed cytoplasmic aggregates, whereas wild-type Crg1-daGFP was distributed normally in the cytoplasm. Interestingly, the Crg1-methyltransferase domain point mutants (D44A, D67A, and E105A/D108A) also showed the same cytoplasmic aggregates as Crg1ΔN-daGFP. In silico prediction of the tertiary structures of these mutants indicated an altered protein conformation. Altogether, these observations suggest that the N-terminal truncation, as well as the point mutations in the methyltransferase domain, alters the native folding of Crg1 methyltransferase, resulting in a loss of enzyme activity. Furthermore, the crg1ΔN mutant showed the same phenotypes as the crg1 null mutant in the presence of cantharidin, i.e., lethal cell growth, PE auxotrophy, temperature sensitivity, endoplasmic reticulum stress, GPI anchor missorting, and cell wall damage. Overall, this study identifies an essential role of the N-terminus of Crg1 in methyltransferase activity and cantharidin resistance.

Previously uncharacterized amino acid residues in histone H3 and H4 mutants with roles in DNA damage repair response and cellular aging.

Chromatin regulates gene expression and genome maintenance, and consists of histones and other components. The post-translational modification of histones plays a key role in maintaining the structure and function of chromatin under different pathophysiological stress conditions. Here, we investigate the functions of previously unexplored amino acid residues in histones H3 and H4. To do so, we screened a library of yeast histone mutants following DNA damage and identified that substitution mutations of histone H3 (H3Q5A/E and H3Q120A) and H4 (H4Y88A/E and H4R78K) render yeast cells sensitive to DNA-damaging agents. These histone mutants show an activated DNA damage response, Rad53 phosphorylation and Sml1 degradation in the presence of methyl methanesulfonate (MMS). In histone H3Q5A/E mutants, RNR2 and RNR3 genes were induced at low level, as was RNR3 in H4 histone mutants following DNA damage. In H3 mutant cells, the cell cycle was deregulated, leading to inefficient cell cycle arrest in the presence of MMS, and genes involved in aging and DNA damage repair pathways were constitutively upregulated. In H3 mutants (H3Q5A, H3Q5E and H3Q120A), we observed reduced chronological lifespan (CLS), compared with extended CLS in the H4R78K mutant. Histone mutants also showed altered H3K4me and H3K56ac modifications and improper activation of the stress responsive Slt2 and Hog1 kinases. Thus, we have determined the significance of previously uncharacterized residues of H3 and H4 in DNA damage response, cell cycle progression and cellular aging.

Casein kinase 2 inhibition impairs spontaneous and oxytocin-induced contractions in late pregnant mouse uterus.

NEW FINDINGS: What is the central question of this study? Does the inhibition of the protein kinase casein kinase 2 (CK2) alter the uterine contractility? What is the main finding and its importance? Inhibition of CK2 impaired the spontaneous and oxytocin-induced contractility in late pregnant mouse uterus. This finding suggests that CK2 is a novel pathway mediating oxytocin-induced contractility in the uterus and thus opens up the possibility for this class of drugs to be developed as a new class of tocolytics. ABSTRACT: The protein kinase casein kinase 2 (CK2) is a ubiquitously expressed serine or threonine kinase known to phosphorylate a number of substrates. The aim of this study was to assess the effect of CK2 inhibition on spontaneous and oxytocin-induced uterine contractions in 19 day pregnant mice. The CK2 inhibitor CX-4945 elicited a concentration-dependent relaxation in late pregnant mouse uterus. CX-4945 and another selective CK2 inhibitor, apigenin, also inhibited the oxytocin-induced contractile response in late pregnant uterine tissue. Apigenin also blunted the prostaglandin F2α response, but CX-4945 did not. Casein kinase 2 was located in the lipid raft fractions of the cell membrane, and disruption of lipid rafts was found to reverse its effect. The results of the present study suggest that CK2, located in lipid rafts of the cell membrane, is an active regulator of spontaneous and oxytocin-induced uterine contractions in the late pregnant mouse.

A systematic assessment of chemical, genetic, and epigenetic factors influencing the activity of anticancer drug KP1019 (FFC14A).

KP1019 ([trans-RuCl4(1H-indazole)2]; FFC14A) is one of the promising ruthenium-based anticancer drugs undergoing clinical trials. Despite the pre-clinical and clinical success of KP1019, the mode of action and various factors capable of modulating its effects are largely unknown. Here, we used transcriptomics and genetic screening approaches in budding yeast model and deciphered various genetic targets and plethora of cellular pathways including cellular signaling, metal homeostasis, vacuolar transport, and lipid homeostasis that are primarily targeted by KP1019. We also demonstrated that KP1019 modulates the effects of TOR (target of rapamycin) signaling pathway and induces accumulation of neutral lipids (lipid droplets) in both yeast and HeLa cells. Interestingly, KP1019-mediated effects were found augmented with metal ions (Al3+/Ca2+/Cd2+/Cu2+/Mn2+/Na+/Zn2+), and neutralized by Fe2+, antioxidants, osmotic stabilizer, and ethanolamine. Additionally, our comprehensive screening of yeast histone H3/H4 mutant library revealed several histone residues that could significantly modulate the KP1019-induced toxicity. Altogether, our findings in both the yeast and HeLa cells provide molecular insights into mechanisms of action of KP1019 and various factors (chemical/genetic/epigenetic) that can alter the therapeutic efficiency of this clinically important anticancer drug.

Effect of Calotropis procera (madar) and amprolium supplementation on parasitological parameters of broilers during mixed Eimeria species infection.

AIM: An experiment was conducted on day old 168 broiler chicks to study the effect of 0.4% as well as 0.2% Calotropis procera (madar) leaf powder and 0.0125% amprolium supplementation on parasitological parameters of broilers during mixed Eimeria species infection. MATERIALS AND METHODS: Chicks were randomly divided into seven groups (I-VII) each with two replicates of 12 chicks. On 15th day of experiment, broilers of Group II, IV, VI, and VII were infected with 50,000 sporulated oocysts of mixed Eimeria species. To evaluate the anticoccidial effect of different feed supplements percent fecal score, percent survival, percent weight gain, performance index (PI), average oocyst production, and percent reduction in oocyst production were calculated. RESULTS: It was observed that amprolium supplementation had maximum anticoccidial effect as it gave the best efficacy in terms of all parameters, whereas supplementation of 0.4% madar leaf powder showed nonsignificant difference with amprolium for some parameters such as percent survival, percent weight gain, and PI. CONCLUSION: It can be concluded that madar (C. procera) leaf powder and amprolium had comparable activity against coccidiosis. Hence, madar leaf powder may be used for the prevention and control of mixed Eimeria spp. infection prevalent in field conditions.

In vitro Histone H3 Cleavage Assay for Yeast and Chicken Liver H3 Protease.

Histone proteins are subjected to a wide array of reversible and irreversible post-translational modifications (PTMs) (Bannister and Kouzarides, 2011; Azad and Tomar, 2014). The PTMs on histones are known to regulate chromatin structure and function. Histones are irreversibly modified by proteolytic clipping of their tail domains. The proteolytic clipping of histone tails is continuously attracting interest of researchers in the field of chromatin biology. We can recapitulate H3-clipping by performing in vitro H3 cleavage assay. Here, we are presenting the detailed protocol to perform in vitro H3 cleavage assay.

Biochemical Analysis Reveals the Multifactorial Mechanism of Histone H3 Clipping by Chicken Liver Histone H3 Protease.

Proteolytic clipping of histone H3 has been identified in many organisms. Despite several studies, the mechanism of clipping, the substrate specificity, and the significance of this poorly understood epigenetic mechanism are not clear. We have previously reported histone H3 specific proteolytic clipping and a protein inhibitor in chicken liver. However, the sites of clipping are still not known very well. In this study, we attempt to identify clipping sites in histone H3 and to determine the mechanism of inhibition by stefin B protein, a cysteine protease inhibitor. By employing site-directed mutagenesis and in vitro biochemical assays, we have identified three distinct clipping sites in recombinant human histone H3 and its variants (H3.1, H3.3, and H3t). However, post-translationally modified histones isolated from chicken liver and Saccharomyces cerevisiae wild-type cells showed different clipping patterns. Clipping of histone H3 N-terminal tail at three sites occurs in a sequential manner. We have further observed that clipping sites are regulated by the structure of the N-terminal tail as well as the globular domain of histone H3. We also have identified the QVVAG region of stefin B protein to be very crucial for inhibition of the protease activity. Altogether, our comprehensive biochemical studies have revealed three distinct clipping sites in histone H3 and their regulation by the structure of histone H3, histone modifications marks, and stefin B.

Efficient expression and purification of biologically active human cystatin proteins.

Cystatins are reversible cysteine protease inhibitor proteins. They are known to play important roles in controlling cathepsins, neurodegenerative disease, and in immune system regulation. Production of recombinant cystatin proteins is important for biochemical and function characterization. In this study, we cloned and expressed human stefin A, stefin B and cystatin C in Escherichia coli. Human stefin A, stefin B and cystatin C were purified from soluble fraction. For cystatin C, we used various chaperone plasmids to make cystatin C soluble, as it is reported to localize in inclusion bodies. Trigger factor, GroES-GroEL, DnaK-DnaJ-GrpE chaperones lead to the presence of cystatin C in the soluble fraction. Immobilized metal affinity chromatography, glutathione sepharose and anion exchange chromatography techniques were employed for efficient purification of these proteins. Their biological activities were tested by inhibition assays against cathepsin L and H3 protease.

H3 clipping activity of glutamate dehydrogenase is regulated by stefin B and chromatin structure.

Glutamate dehydrogenase has been recently identified as a tissue-specific histone H3-specific clipping enzyme. We have previously shown that it cleaves free as well as chromatin-bound histone H3. However, the physiological significance of this enzyme is still not clear. The present study aimed to improve our understanding of its significance in vivo. Using biochemical and cell biological approaches, we show that glutamate dehydrogenase is primarily associated with euchromatin, and it re-localizes from the nuclear periphery to the nucleolus upon DNA damage. The cysteine protease inhibitor stefin B regulates the H3 clipping activity of the enzyme. Chromatin structure and certain histone modifications influence H3 clipping activity. Interestingly, we also observed that an in vivo truncated form of H3 lacks H3K56 acetylation, which is a code for the DNA damage response. Together, these results suggest that glutamate dehydrogenase is a euchromatin-associated enzyme, and its H3 clipping activity is regulated by chromatin structure, histone modifications and an in vivo inhibitor. In response to DNA damage, it re-localizes to the nuclei, and hence may be involved in regulation of gene expression in vivo.