Combined Density Functional Theory (DFT) and Continuum Calculations of pK(a) in Carbonic Anhydrase

Deprotonation of zinc-bound water in carbonic anhydrase II is the rate-limiting step in the catalysis of carbon dioxide between gas- and water-soluble forms. To understand the factors determining the extent of dissociation, or pK(a), of the zinc-bound water, we apply quantum chemistry calculations to the active site coupled with a continuum model of the surrounding environment. Experimentally determined changes in pK(a) associated with mutations of the active site are well reproduced by this approach. Analysis of the active site structure and charge/dipole values provides evidence that mutations cause changes in both conformation of the active site structure and local polarization, which accounts for the shifts in pK(a). More specifically, the shifts in pK(a) correlate with the dipole moments of the zinc-bound water upon deprotonation. The data further support the conclusion that the distinct pK(a) values found in mutations of the same type, but applied to different sites, result from asymmetric ligation and different electronic environments around the zinc ion.     

Altered Levels of Trace Elements in Acute Lung Injury After Severe Trauma

Trace element (TE) supplementation can reduce the incidence of multiple organ failure after severe trauma. The lung plays a main role in post-injury multiple organ failure. In the present study, the relationship between TEs and acute lung injury (ALI) post-injury was investigated in a rabbit model of severe trauma with an injury severity score of 27. New Zealand white rabbits were randomly assigned to trauma-control, trauma-TE groups, and a control group. During days 1-5 post-trauma, each rabbit in the trauma-TE group received 0.1 ml multi-TE compound intraperitoneally to give a daily dose of 32.50 mg/kg of Zn, 6.35 mg/kg of Cu, 1.38 mg/kg of Mn, and 0.16 mg/kg of Se. Concentrations of blood and lung selenium (Se), copper (Cu), zinc (Zn), and manganese (Mn) were measured at 6 and 24 h, as well as 3, 6, 9, and 14 days after trauma. Levels of glutathione peroxidase (GPx), total superoxide dismutase (SOD), Cu/Zn superoxide dismutase (Cu/Zn-SOD), and malondialdehyde (MDA) in serum and lung tissue and the level of intercellular adhesion molecular-1 (ICAM-1) in serum were detected simultaneously. In addition, the lung coefficient (LC) and the lung permeation index (LPI) were measured. Serum and lung Zn, Se, and Mn levels decreased dramatically by 6 h after trauma in both experimental groups. Cu showed no significant changes after trauma. The serum and lung GPx and SOD levels in the experimental group decreased significantly on days 1 and 3, respectively. Serum and lung MDA began to increase on day 3 in the trauma group but increased less after TE supplementation. Serum ICAM-1 peaked on day 6 in the experimental group. LC and LPI increased gradually post-trauma, peaking on days 6 and 9, respectively. In conclusion, an acute lung injury causes declines of the levels of TEs in serum and lung which can be significantly prevented by TE supplementation and which can also mitigate some of the morphological and biomechanical changes in ALI.     

Molecular characteristics of Camallanus Spp. (Spirurida: Camallanidae) in fishes from China based on its rDNA sequences

In the paper, we explored the intra- and interspecific evolutionary variation among species of Camallanus collected from different fish species in various regions of China. We determined the internal transcribed spacers of ribosomal DNA (ITS rDNA) sequences of these nematodes. The divergence (uncorrected p-distance) of ITS1, ITS2, and ITS rDNA data sets confirmed 2 valid species of Camallanus in China, i.e., C. cotti and C. hypophthalmichthys. The 2 species were distinguished not only by their different morphologies and host ranges but also by a tetranucleotide microsatellite (TTGC)n present in the ITS1 region of C. cotti. Phylogenetic analyses of the nematodes disclosed 2 main clades, corresponding to different individuals of C. cotti and C. hypophthalmichthys from different fish species in various geographical locations, although the interior nodes of each clade received poor support.     

A new species of Allocreadium (Trematoda: Allocreadiidae) from freshwater fishes in the Danjiangkou Reservoir in China

A new species of Allocreadium, Allocreadium danjiangensis n. sp., is described from the intestine of several species of freshwater fish, including Abbottina rivularis (Basilewsky, 1855), Sarcocheilichthys nigripinnis nigripinns (Güther, 1873), Gnathopogon argentatus (Sauvage et Dabry 1874), Opsariichthys uncirostris bidens (Gunther, 1873), and Erythroculter mongolicus mongolicus (Basilewsky, 1855) (Cyprinidae) from the Danjiangkou Reservoir in central China. The main morphological characters of the new species are as follows: vitelline follicles numerous, extending from the level of acetabulum to posterior extremity, distributed over both sides around the ceca; cirrus sac relatively large, developed, lying obliquely anterior to the acetabulum, extending from the level of the intestinal bifurcation to the central level of acetabulum, and overlapping left or right cecal; and ovary much smaller than testes, generally close to or even overlapping the anterior border of anterior testis. Observation by scanning electron microscopy shows only 2 kinds of tegumental formations, i.e., papillae and tubercles, instead of 3 types of tegumental formations, i.e., papillae, bosses, and minute sensor receptors observed on other species of the Allocreadiidae. The tegumental striations of the present species vary on the different parts of the body. In addition, a new structure, identified as the "groove" with a tonguelike tubercle, was observed on the inner wall of acetabulum.     

Structural mechanism for bacterial oxidation of oceanic trimethylamine into trimethylamine N‐oxide

Trimethylamine (TMA) and trimethylamine N‐oxide (TMAO) are widespread in the ocean and are important nitrogen source for bacteria. TMA monooxygenase (Tmm), a bacterial flavin‐containing monooxygenase (FMO), is found widespread in marine bacteria and is responsible for converting TMA to TMAO. However, the molecular mechanism of TMA oxygenation by Tmm has not been explained. Here, we determined the crystal structures of two reaction intermediates of a marine bacterial Tmm (RnTmm) and elucidated the catalytic mechanism of TMA oxidation by RnTmm. The catalytic process of Tmm consists of a reductive half‐reaction and an oxidative half‐reaction. In the reductive half‐reaction, FAD is reduced and a C4a‐hydroperoxyflavin intermediate forms. In the oxidative half‐reaction, this intermediate attracts TMA through electronic interactions. After TMA binding, NADP⁺ bends and interacts with D317, shutting off the entrance to create a protected micro‐environment for catalysis and exposing C4a‐hydroperoxyflavin to TMA for oxidation. Sequence analysis suggests that the proposed catalytic mechanism is common for bacterial Tmms. These findings reveal the catalytic process of TMA oxidation by marine bacterial Tmm and first show that NADP⁺ undergoes a conformational change in the oxidative half‐reaction of FMOs.     

Ligand binding and conformational changes of SUR1 subunit in pancreatic ATP-sensitive potassium channels

ATP-sensitive potassium channels (K_ATP) are energy sensors on the plasma membrane. By sensing the intracellular ADP/ATP ratio of β-cells, pancreatic K_ATP channels control insulin release and regulate metabolism at the whole body level. They are implicated in many metabolic disorders and diseases and are therefore important drug targets. Here, we present three structures of pancreatic K_ATP channels solved by cryoelectron microscopy (cryo-EM), at resolutions ranging from 4.1 to 4.5 Å. These structures depict the binding site of the antidiabetic drug glibenclamide, indicate how Kir6.2 (inward-rectifying potassium channel 6.2) N-terminus participates in the coupling between the peripheral SUR1 (sulfonylurea receptor 1) subunit and the central Kir6.2 channel, reveal the binding mode of activating nucleotides, and suggest the mechanism of how Mg-ADP binding on nucleotide binding domains (NBDs) drives a conformational change of the SUR1 subunit.     

分蘖洋葱试管鳞茎微繁技术研究

以MS+BA 0.4 mg/L+NAA 0.1 mg/L为起始培养基,将获得的分蘖洋葱丛生芽进行试管微鳞茎诱导实验,结果表明：离体条件下,温度和光周期对鳞茎的形成起着决定性的作用, 提高培养基的pH值、增加蔗糖浓度对试管苗鳞茎化也有重要的作用。