Ultrapotent neutralizing antibodies against SARS-CoV-2 with a high degree of mutation resistance.

Many SARS-CoV-2 neutralizing antibodies (nAbs) lose potency against variants of concern. In this study, we developed 2 strategies to produce mutation-resistant antibodies. First, a yeast library expressing mutant receptor binding domains (RBDs) of the spike protein was utilized to screen for potent nAbs that are least susceptible to viral escape. Among the candidate antibodies, P5-22 displayed ultrahigh potency for virus neutralization as well as an outstanding mutation resistance profile. Additionally, P14-44 and P15-16 were recognized as mutation-resistant antibodies with broad betacoronavirus neutralization properties. P15-16 has only 1 binding hotspot, which is K378 in the RBD of SARS-CoV-2. The crystal structure of the P5-22, P14-44, and RBD ternary complex clarified the unique mechanisms that underlie the excellent mutation resistance profiles of these antibodies. Secondly, polymeric IgG enhanced antibody avidity by eliminating P5-22's only hotspot, residue F486 in the RBD, thereby potently blocking cell entry by mutant viruses. Structural and functional analyses of antibodies screened using both potency assays and the yeast RBD library revealed rare, ultrapotent, mutation-resistant nAbs against SARS-CoV-2.

The role of satellite glial cells in orofacial pain.

Some chronic pain conditions in the orofacial region are common, the mechanisms underlying which are unresolved. Satellite glial cells (SGCs) are the glial cells of the peripheral nervous system. In the sensory ganglia, each neuronal body is surrounded by SGCs forming distinct functional units. The unique structural organization enables SGCs to communicate with each other and with their enwrapped neurons via a variety of ways. There is a growing body of evidence that SGCs can influence the level of neuronal excitability and are involved in the development and/or maintenance of pain. The aim of this review was to summarize the latest advances made about the implication of SGCs in orofacial pain. It may offer new targets for the development of orofacial pain treatment.

Cause of death in Chinese Takayasu arteritis patients.

To analyze the causes of death and the related risk factors for in-patients with Takayasu arteritis (TAK) admitted to a referral center of China during 1983 to 2014.The medical charts of 12 deceased TAK patients (10 women, 2 men) were reviewed by two senior rheumatologists. The demographic data, clinical manifestations, angiographic presentations, and the direct causes of death were analyzed retrospectively. Medical records of 40 TAK patients (32 women, 8 men) were selected as controls by age and sex matching method from 81 patients who were sampled isometrically from 810 successively admitted TAK in-patients of the same center during the same period. In addition to the comparison of clinical manifestations between the two groups, binary logistic regression was conducted to explore the related risk factors of mortality of TAK.Twelve patients died at the median age of 33.5 (ranging from 13 to 68 years old). The median survival time was 102.5(ranging from 6 to 567) months. The direct causes of death were heart failure in 5 (5/12, 41.7%), hemorrhage in 2 (2/12, 16.7%), pulmonary infection in 2 (2/12, 16.7%), sudden death in 1 (1/12, 8.3%), postoperative complication in 1 (1/12, 8.3%), and end-stage malignancy in 1 (1/12, 8.3%). Ischemia (4/12, 33.3%) and hemorrhage (4/12, 33.3%) were the two most common presentations in deceased patients. Eight patients had received surgical procedures related to TAK changes. Among them, 2 patients died after surgical procedure, the other 6 patients died later of non-operation-related causes. Compared with the control group (n = 40), patients in the deceased group had longer disease duration (P = 0.017), higher proportion of active disease (P = 0.020), secondary hypertension (P = 0.004), and congestive heart failure (P = 0.017). A model of binary logistic regression had revealed that secondary hypertension (odds ratio [OR] = 9.333, 95% confidence interval [CI]: 1.721 - 50.614, P = 0.010), congestive heart failure (OR = 5.667, 95% CI: 1.248 - 25.734, P = 0.025), and longer disease duration (OR = 1.007, 95% CI: 1.001 - 0.735, P = 0.027) were risk factors for TAK mortality. Active disease (OR = 0.167, 95% CI: 0.038 - 50.614, P = 0.018) was negatively associated with death of TAK.Heart failure is the leading cause of death in TAK patients, followed by ischemia and pulmonary infection. Early deaths occur postoperatively but become rare later after the procedure. Well-control of hypertension, and prevention of congestive heart failure may improve the long-term prognosis.

MicroRNA let-7i induced autophagy to protect T cell from apoptosis by targeting IGF1R.

MicroRNA let-7i is up-regulated in T cells from patients with Ankylosing Spondylitis (AS). In this study, we investigated the role of let-7i in T cells survival. Our results demonstrated down-regulation of insulin-like growth factor-1 receptor (IGF1R) in T cells from patients with AS. Luciferase reporter assay suggested IGF1R as direct target of let-7i. Overexpression of let-7i in Jurkat cells significantly suppressed IGF1R expression, which mimicked the action of IGF1R siRNA. IGF1R inhibition led to a strinking decrease in phosphorylation of mTOR and Akt, down-regulation of Bcl-2, up-regulation of Bax and cleavage of caspase 3 and PARP. Meanwhile, IGF1R inhibition induced autophagy. Autophagy induced by let-7i overexpression contributed to protect cells from apoptosis. Our data indicated that let-7i might control T cells fates in AS by targeting IGF1R.

Engineered cytidine triphosphate synthetase with reduced product inhibition.

Cytidine triphosphate (CTP) synthetase (CTPS) (EC number 6.3.4.2) is a key enzyme involved in de novo synthesis of CTP. It catalyzes the rate-limiting step of the process due to the product inhibition effects on the enzyme. In this study, a novel CTPS from Corynebacterium glutamicum ATCC 13032 (CgCTPS) was cloned, expressed and characterized. A series of mutagenesis in its N-terminal ammonia ligase (ALase) domain was performed in order to reduce CTP product inhibition. All single mutation variants (D160E, E162A, E168K) lowered product inhibition by lowering the enzyme's binding affinity for CTP. The homology model of CgCTPS showed that D160E mutant caused steric hindrance for the pyrimidine ring of CTP stacking, E162A disrupted the hydrogen bond between CTP ribose and side chain and D168K caused minor localized structure perturbations of CTP binding pocket. The triple mutant of CTPS (D160E-E162A-E168K) with halved Km, doubled Vmax and the 23.5-fold increased IC50 for CTP shows a potential for use in industrial-scale CTP production by its better performance in enzyme kinetics and product inhibition.

Gene cloning, expression, and characterization of a cyclic nucleotide phosphodiesterase from Arthrobacter sp. CGMCC 3584.

Based on thermal asymmetric interlaced polymerase chain reaction, the arpde gene encoding a cyclic nucleotide-specific phosphodiesterase was cloned from Arthrobacter sp. CGMCC 3584 for the first time. The 930-bp region encoded a 309-amino-acid protein with a molecular weight of 33.6 kDa. The recombinant ArPDE was able to hydrolyze 3',5'-cAMP, 3',5'-cGMP, and 2',3'-cAMP. The K m values of ArPDE for 3',5'-cAMP and 3',5'-cGMP were 6.82 and 12.82 mM, respectively. ArPDE was thermostable and displayed optimal activity at 45 °C and pH 7.5. The enzyme did not require any metal cofactors, although its activity was stimulated by 2 mM Co(2+) and inhibited by Zn(2+). Nucleotides, reducing agents, and sulfhydryl reagents had different inhibitory effects on the activity of ArPDE. NaF, the actual compound used to improve the industrial yield of cAMP, exhibited 62 % inhibitions at concentrations of 10 mM.