ABSTRACT
BACKGROUND: Ferredoxin NADP(H) oxidoreductases (EC 1.18.1.2) (FNR) are flavoenzymes present in photosynthetic organisms; they are relevant for the production of reduced donors to redox reactions, i.e. in photosynthesis, the reduction of NADP+ to NADPH using the electrons provided by Ferredoxin (Fd), a small FeS soluble protein acceptor of electrons from PSI in chloroplasts. In rhodophyta no information about this system has been reported, this work is a contribution to the molecular and functional characterization of FNR from Gracilaria chilensis, also providing a structural analysis of the complex FNR/Fd. METHODS: The biochemical and kinetic characterization of FNR was performed from the enzyme purified from phycobilisomes enriched fractions. The sequence of the gene that codifies for the enzyme, was obtained using primers designed by comparison with sequences of Synechocystis and EST from Gracilaria. 5'RACE was used to confirm the absence of a CpcD domain in FNRPBS of Gracilaria chilensis. A three dimensional model for FNR and Fd, was built by comparative modeling and a model for the complex FNR: Fd by docking. RESULTS: The kinetic analysis shows KMNADPH of 12.5 M and a kcat of 86 s-1, data consistent with the parameters determined for the enzyme purified from a soluble extract. The sequence for FNR was obtained and translated to a protein of 33646 Da. A FAD and a NADP+ binding domain were clearly identified by sequence analysis as well as a chloroplast signal sequence. Phycobilisome binding domain, present in some cyanobacteria was absent. Transcriptome analysis of Gch revealed the presence of two Fd; FdL and FdS, sharing the motif CX5CX2CX29X. The analysis indicated that the most probable partner for FNR is FdS. CONCLUSION: The interaction model produced, was consistent with functional properties reported for FNR in plants leaves, and opens the possibilities for research in other rhodophyta of commercial interest.
Subject(s)
Gracilaria/enzymology , Ferredoxin-NADP Reductase/chemistry , Ferredoxins/metabolism , Oxidation-Reduction , Photosynthesis/physiology , Amino Acid Sequence , Gracilaria/chemistry , Electrophoresis, Polyacrylamide Gel , Ferredoxin-NADP Reductase/genetics , Ferredoxin-NADP Reductase/pharmacokineticsABSTRACT
The human pregnane X receptor (hPXR) plays a critical role in the metabolism, transport and clearance of xenobiotics in the liver and intestine. The hPXR can be activated by a structurally diverse of drugs to initiate clinically relevant drug-drug interactions. In this article, in silico investigation was performed on a structurally diverse set of drugs to identify critical structural features greatly related to their agonist activity towards hPXR. Heuristic method (HM)-Best Subset Modeling (BSM) and HM-Polynomial Neural Networks (PNN) were utilized to develop the linear and non-linear quantitative structure-activity relationship models. The applicability domain (AD) of the models was assessed by Williams plot. Statistically reliable models with good predictive power and explain were achieved (for HM-BSM, r (2)=0.881, q LOO (2) =0.797, q EXT (2) =0.674; for HM-PNN, r (2)=0.882, q LOO (2) =0.856, q EXT (2) =0.655). The developed models indicated that molecular aromatic and electric property, molecular weight and complexity may govern agonist activity of a structurally diverse set of drugs to hPXR.
Subject(s)
Humans , Computer Simulation , Models, Statistical , Molecular Weight , Neural Networks, Computer , Quantitative Structure-Activity Relationship , Receptors, Steroid , Chemistry , Small Molecule Libraries , Chemistry , Static ElectricityABSTRACT
Objective To study the ultra-structural features of interstitial cajal cells (ICC)in fetal enteron and then classify them.Methods Ultra-structural features of ICC in fetal enteron with spontaneous abortion or premature birth were detected under JEM-2000EX transmission electron microscope for the cause of fetal death, including two fetus specimens of a gestation 18 months and 28 months stained with lead nitrate and uranium acetate.Results ICC had a big oval karyon and a little of cytoplasm. ICC formed spindle or stellate cells with 2 to 5 long cell processes . From the esophagus to the terminal ileum ICC mostly had the same ultra-structural features, but with mitochondria and a well-developed endoplasmic reticulum and Golgi complex in the whole colon and the rectum , more than those of esophagus and small intestine. In the whole colon and rectum ICC had similar ultra-structural features. ICC also possessed an abundance of mitochondria and a well-developed endoplasmic reticulum and Golgi complex. ICC also possessed Caveloae lipid droplet with more electron dense and heterochromatin. Two types of ICC were identified under JEM-2000EX transmission electron microscope. One type was present from the esophagus to the terminal ileum and the other type was observed in the colon and rectum. The first type ICC in circular muscle layer was bipolar cells which extended to tapering processes in opposite directions. These processes rarely branched, and their appearance was similar to smooth muscle cells.Most of them ran parallel to the circular muscles . The second type of ICC in the myenteric plexus and longitudinal muscle layer was mostly multipolar and rarely bipolar cells with long processes. They showed an irregular appearance characterized by numerous short spike-like branches. Processes of multipolar cells extended in every direction and connected with each other. ICC nerve cells and smooth muscle cells were connected with gap-like junction, which was the main connection mode .Conclusions The ultra-structural features of ICC in fetal enteron varied with the diffe-rent locus and different tissue sheets in the enteron. The mitochondria and a well-developed endoplasmic reticulum and Golgi complex of the whole colon and rectum are more and more developed than that of esophagus and small intestine. Ultra-structural features of ICC will develop further with the gestational age. The gap- like junction among ICC nerve cell and smooth muscle cell are highly important for ICC to educe function.
ABSTRACT
AIM: To characterize the chemical structure of a homogenous polysaccharide MP-3b obtained from Mulberry leaves. METHODS: Sugar component analysis,methylation analysis,partial acid hydrolysis,IR,ESI-MS,NMR were applied to determine the structural features. RESULTS: The molecular weight of MP-3b was 8.9?10~4 dalton by HPGFC.MP-3b consisted of rhammose,arabinose,xylose,glucose,galactose and galacturonic acid units in the molar ratio of 0.65:1.00:0.23:0.42:0.98:0.60.The chain of MP-3b included terminal,1,5-linked,1,3,5-linked Ara and terminal,1,2-linked,1,2,4-linked Rha,and 1,4-linked Xyl,and terminal Glc,and terminal,1,3-linked,1,6-linked,1,3,6-linked Gal and 1,4-linked,1,3,4-linked GalA.The side chain contained branched(1→5) linked arabinan. CONCLUSION: MP-3b is a new acidic polysaccharide obtained from Mulberry leaves for the first time.