Thermal aggregation of human immunoglobulin G in arginine solutions: Contrasting effects of stabilizers and destabilizers

Arginine is widely used as aggregation suppressor of proteins in biotechnology and pharmaceutics. However, why the effect of arginine depends on the types of proteins and stresses, including monoclonal antibodies, is still unclear. Here we investigated the precise processes of the thermal aggregation of human immunoglobulin G (IgG) in the presence of additives. As expected, arginine was the best additive to suppress the formation of insoluble aggregates during heat treatment, though it was unable to preserve the monomer content. A systematic analysis of the additives showed that sugars and kosmotropic ion inhibit the formation of soluble oligomers. These behaviors indicate that the thermal aggregation of IgG occurs by (i) the formation of soluble oligomers, which is triggered by the unfolding process that can be stabilized by typical osmolytes, and (ii) the formation of insoluble aggregates through weak cluster–cluster interactions, which can be suppressed by arginine. Understanding the detailed mechanism of arginine will provide useful information for the rational formulation design of antibodies

Antiviral and Virucidal Activities of Nα-Cocoyl-L-Arginine Ethyl Ester

Various amino acid-derived compounds, for example, Nα-Cocoyl-L-arginine ethyl ester (CAE), alkyloxyhydroxylpropylarginine, arginine cocoate, and cocoyl glycine potassium salt (Amilite), were examined for their virucidal activities against herpes simplex virus type 1 and 2 (HSV-1 and HSV-2), influenza A virus (IAV), and poliovirus type 1 (PV-1) in comparison to benzalkonium chloride (BKC) and sodium dodecylsulfate (SDS) as a cationic and anionic control detergent and also to other commercially available disinfectants. While these amino acid-derived compounds were all effective against HSV-1 and HSV-2, CAE and Amilite were the most effective. These two compounds were, however, not as effective against IAV, another enveloped virus, as against HSV. Cytotoxicity of CAE was weak; at 0.012%, only 5% of the cells were killed under the conditions, in which 100% cells were killed by either SDS or BKC. In addition to these direct virucidal effects, CAE inhibited the virus growth in the HSV-1- or PV-1-infected cells even at 0.01%. These results suggest a potential application of CAE as a therapeutic or preventive medicine against HSV superficial infection at body surface

A single Gly114Arg mutation stabilizes the hexameric subunit assembly and changes the substrate specificity of halo-archaeal nucleoside diphosphate kinase

AbstractNucleoside diphosphate kinase from extremely halophilic archaeon (HsNDK) requires above 2M NaCl concentration for in vitro refolding. Here an attempt was made to isolate mutations that allow HsNDK to refold in low salt media. Such a screening resulted in isolation of an HsNDK mutant, Gly114Arg, which efficiently refolded in the presence of 1M NaCl. This mutant, unlike the wild type enzyme, was expressed in Escherichia coli as an active form. The residue 114 is in close proximity to Glu155 of the neighboring subunit in the three dimensional hexameric structure of the HsNDK. It is thus possible that the attractive electrostatic interactions occur between Arg114 and Glu155 in the mutant HsNDK, stabilizing the hexameric subunit assembly

Arginine and lysine reduce the high viscosity of serum albumin solutions for pharmaceutical injection

Therapeutic protein solutions for subcutaneous injection must be very highly concentrated, which increases their viscosity through protein–protein interactions. However, maintaining a solution viscosity below 50 cP is important for the preparation and injection of therapeutic protein solutions. In this study, we examined the effect of various amino acids on the solution viscosity of very highly concentrated bovine serum albumin (BSA) and human serum albumin (HSA) at a physiological pH. Among the amino acids tested, l-arginine hydrochloride (ArgHCl) and l-lysine hydrochloride (LysHCl) (50–200 mM) successfully reduced the viscosity of both BSA and HSA solutions; guanidine hydrochloride (GdnHCl), NaCl, and other sodium salts were equally as effective, indicating the electrostatic shielding effect of these additives. Fourier transform infrared spectroscopy showed that BSA is in its native state even in the presence of ArgHCl, LysHCl, and NaCl at high protein concentrations. These results indicate that weakened protein–protein interactions play a key role in reducing solution viscosity. ArgHCl and LysHCl, which are also non-toxic compounds, will be used as additives to reduce the solution viscosity of concentrated therapeutic proteins

Distribution of seawater turbidity due to dissolved organic matter and suspended matter in Tokyo Bay and the correlation with the contributing matter

東京海洋大学海洋科学部海洋環境学科東京海洋大学海洋科学部海洋環境学科西海区研究所石垣支所海洋環境研究室東京海洋大学海洋科学部海洋環境学

Grading Meningioma : A Comparative Study of Thallium-SPECT and FDG-PET

The purpose was to compare capability of fluorine-18 fluorodeoxyglucose (FDG)-PET and thallium-201 (Tl)-SPECT for grading meningioma. This retrospective study was conducted as a case-control study under approval by the institutional review board. In the hospital information system, 67 patients (22 men and 45 women) who had both FDG-PET and Tl-SPECT preoperative examinations were found with histopathologic diagnosis of meningioma. The maximum FDG uptake values of the tumors were measured, and they were standardized to the whole body (SUVmax) and normalized as gray matter ratio (SUVRmax). Mean and maximum Tl uptake ratios (TURmean and TURmax, respectively) of the tumors were measured and normalized as ratios to those of the contralateral normal brain. Receiver-operating characteristic curve analyses of the 4 indexes were conducted for differentiation between low- and high-grade meningiomas, and areas under the curves (AUCs) were compared. Correlation coefficients were calculated between these indexes and Ki-67. Fifty-six meningiomas were classified as grade I (low grade), and 11 were grade II or III (high grade). In all 4 indexes, a significant difference was observed between low- and high-grade meningiomas (P<0.05). AUCs were 0.817 (SUVmax), 0.781 (SUVRmax), 0.810 (TURmean), and 0.831 (TURmax), and no significant difference was observed among the indexes. Their sensitivity and specificity were 72.7% to 90.9% and 71.4% to 87.5%, respectively. Correlation of the 4 indexes to Ki-67 was statistically significant, but coefficients were relatively low (0.273-0.355). Tl-SPECT, which can be used at hospitals without a cyclotron or an FDG distribution network, has high diagnostic capability of meningioma grades comparable to FDG-PET

Pillar[6]arene acts as a biosensor for quantitative detection of a vitamin metabolite in crude biological samples

ビタミン代謝物を迅速定量できる超分子バイオセンサーを開発. 京都大学プレスリリース. 2020-12-09.Metabolic syndrome is associated with obesity, hypertension, and dyslipidemia, and increased cardiovascular risk. Therefore, quick and accurate measurements of specific metabolites are critical for diagnosis; however, detection methods are limited. Here we describe the synthesis of pillar[n]arenes to target 1-methylnicotinamide (1-MNA), which is one metabolite of vitamin B3 (nicotinamide) produced by the cancer-associated nicotinamide N-methyltransferase (NNMT). We found that water-soluble pillar[5]arene (P5A) forms host–guest complexes with both 1-MNA and nicotinamide, and water-soluble pillar[6]arene (P6A) selectively binds to 1-MNA at the micromolar level. P6A can be used as a “turn-off sensor” by photoinduced electron transfer (detection limit is 4.38 × 10−6 M). In our cell-free reaction, P6A is used to quantitatively monitor the activity of NNMT. Moreover, studies using NNMT-deficient mice reveal that P6A exclusively binds to 1-MNA in crude urinary samples. Our findings demonstrate that P6A can be used as a biosensor to quantify 1-MNA in crude biological samples

ESI-MS Analysis of Tetrapyridinium Macrocycle Complexation with Carboxylic Anions

Complex formations of a quaternary tetrapyridinium macrocycle (Cpy4Br4) with anionic species, mono-, di-, tri-carboxylic acids, adenocine 5′-triphosphate (ATP), diphosphate (ADP), and monophosphate (AMP) were observed in a neutral pH region. Electrospray ionization mass spectrometry (ESI-MS) was used to study reactivity and selectivity of the complex formations between the Cpy4 cation and the guest anions

A tribute to Dr. Serge N. Timasheff, our mentor

63 p.-6 fig.Dr. Serge N. Timasheff, our mentor and friend, passed away in 2019. This article is a collection of tributes from his postdoctoral fellows, friends, and daughter, who all have been associated with or influenced by him or his research. Dr. Timasheff is a pioneer of research on thermodynamic linkage between ligand interaction and macromolecular reaction. We all learned a great deal from Dr. Timasheff, not only about science but also about life.Peer reviewe