Mutations Increasing Cofactor Affinity, Improve Stability and Activity of a Baeyer-Villiger Monooxygenase.

The typically low thermodynamic and kinetic stability of enzymes is a bottleneck for their application in industrial synthesis. Baeyer-Villiger monooxygenases, which oxidize ketones to lactones using aerial oxygen, among other activities, suffer particularly from these instabilities. Previous efforts in protein engineering have increased thermodynamic stability but at the price of decreased activity. Here, we solved this trade-off by introducing mutations in a cyclohexanone monooxygenase from Acinetobacter sp., guided by a combination of rational and structure-guided consensus approaches. We developed variants with improved activity (1.5- to 2.5-fold) and increased thermodynamic (+5 °C T m) and kinetic stability (8-fold). Our analysis revealed a crucial position in the cofactor binding domain, responsible for an 11-fold increase in affinity to the flavin cofactor, and explained using MD simulations. This gain in affinity was compatible with other mutations. While our study focused on a particular model enzyme, previous studies indicate that these findings are plausibly applicable to other BVMOs, and possibly to other flavin-dependent monooxygenases. These new design principles can inform the development of industrially robust, flavin-dependent biocatalysts for various oxidations.

Rapid antibody diagnostics for SARS-CoV-2 adaptive immune response.

The emergence of a pandemic scale respiratory illness (COVID-19: coronavirus disease 2019) and the lack of the world's readiness to prevent its spread resulted in an unprecedented rise of biomedical diagnostic industries, as they took lead to provide efficient diagnostic solutions for COVID-19. However, these circumstances also led to numerous emergency use authorizations without appropriate evaluation that compromised standards, which could result in a larger than usual number of false-positive or false-negative results, leading to unwanted ambiguity in already confusing realities of the pandemic-hit closures of the world economy. This review is aimed at comparing the claimed or reported clinical sensitivity and clinical specificity of commercially available rapid antibody diagnostics with independently evaluated clinical performance results of the tests. Thereby, we not only present the types of modern antibody diagnostics and their working principles but summarize their experimental evaluations and observed clinical efficiencies to highlight the research, development, and commercialization issues with future challenges. Still, it must be emphasized that the serological or antibody tests do not serve the purpose of early diagnosis but are more suitable for epidemiology and screening populaces with an active immune response, recognizing convalescent plasma donors, and determining vaccine efficacy.

CARLo-7-A plausible biomarker for bladder cancer.

Cancer is defined as undifferentiated and unchecked growth of cells damaging the surrounding tissue. Cancers manifest altered gene expression. Gene expression is regulated by a diverse array of non-protein-coding RNA. Aberrant expression of long non-coding RNAs (lncRNAs) has been recently found to have functional consequences in cancers. In the current study, we report CARLo-7 as the only bladder cancer-specific lncRNA from the CARLos cluster. The expression of this lncRNA correlates with bladder cancer grade. We propose that CARLo-7 has an oncogenic potential and might be regulator of cell proliferation. Furthermore, by comparison the expression of proto-oncogene MYC, which is the only well-annotated gene close to the cancer - associated linkage disequilibrium blocks of this region, does not show a pronounced change in expression between the low- and high-grade tumours. Our results indicate that CARlo-7 can act as a prognostic marker for bladder cancer.

Polymorphism in the IL-8 Gene Promoter and the Risk of Acne Vulgaris in a Pakistani Population.

Interleukin-8 (IL-8) is a well-known inflammatory chemokine and suggested to be involved in the development of acne vulgaris. This study investigates IL-8 plasma levels in acne patients and healthy controls and the molecular basis for the regulation of the IL-8 gene in a Pakistani population. Patients with acne vulgaris (n = 264) and healthy individuals (n = 264) were enrolled in this investigation. Plasma IL-8 levels were determined by enzyme-linked immunosorbent assay (ELISA). The genotyping for IL-8 gene was performed by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP). Our data showed a statistically significant increase in IL-8 levels from acne patients compared with healthy subjects (154.2 ± 52.1 pg/mL in patients vs. 101.6 ± 33.5 pg/mL in controls, p<0.0001). The IL-8-251T>A (rs4073) polymorphism was significantly higher in patients with acne compared with the control group (p=0.013). There was a significant difference between the T and A alleles from acne cases and controls (odds ratio OR=1.6,95 % CI= 1.16-2.19, p=0.003). Logistic-regression analysis showed that the increased IL-8 levels, and the IL-8-251T>A polymorphism were significantly associated with acne. Our data suggest that the elevated IL-8 levels and the IL-8-251T>A polymorphism may be associated with acne vulgaris in the study population.