西双版纳热带次生林中的丛枝菌根调查

对西双版纳热带次生林中13个科的26种植物根系的丛枝菌根真菌（Arbuscular mycorrhizal fongii,AMF）侵染情况进行了研究,并从这些植物的根围土壤中分离鉴定了隶属于球囊霉属（Clomus）、巨孢囊霉属（Gigaspora）、盾巨孢囊霉属（Scutellospora）和无梗囊霉属（Acaulospora）的11种丛枝菌根真菌。该地次生林中AMF的孢子密度为13—29个／100g土壤,平均为19个;种的丰富度在4．9之间（平均为6）;平均频度为53．8％;相对多度为3．2％-26．5％;物种多样性指数和均匀度指数分别为0．94和0．93。丛枝菌根的侵染率达到44．8％．57．2％（平均为50．9％）;球囊霉属（Glomus）和无梗囊霉属（Acattlospora）是热带次生林根围土壤中菌根真菌的优势类群。     

Inhibiting TNF-mediated signaling: a novel therapeutic paradigm for androgen independent prostate cancer

The tumor necrosis factor (TNF) receptor super family comprises of members that induce two distinct signaling cascades, leading to either cell survival or apoptosis. However, in prostate cancer (PCa), TNF-mediated prosurvival signaling is the predominant pathway that leads to cell survival and resistance to therapy. Although inhibition of TNF signaling by pharmacological agents or monoclonal antibodies has gained importance in the field of cancer therapy, toxicity to normal cells has impaired their extensive use for cancer treatment. We previously identified a natural, nontoxic compound psoralidin that inhibited viability and induced apoptosis in androgen independent prostate cancer (AIPC) cells. Thus, the goal of our study is to investigate whether psoralidin inhibits TNF-mediated prosurvival signaling in AIPC cells. Our results suggest that psoralidin inhibits constitutive and TNF-induced expression of TNF-α and its downstream prosurvival signaling molecules such as NF-κB and Bcl-2 in AIPC cells. On the other hand, psoralidin simultaneously induces the death receptor (DR)-mediated apoptotic signaling eventually causing the activation of caspase cascade and resultant induction of apoptosis. Oral administration of psoralidin inhibits expression of TNF-α and NF-κB/p65 in tumor sections, resulting in tumor regression in PC-3 xenografts. Our results suggest that psoralidin inhibits TNF-mediated survival signaling in AIPC and thus is a potent therapeutic agent for prostate cancer.     

Investigating the dynamic properties of the transmembrane segment of phospholamban incorporated into phospholipid bilayers utilizing 2H and 15N solid-state NMR spectroscopy

(2)H and (15)N solid-state NMR spectroscopic techniques were used to investigate the membrane composition, orientation, and side-chain dynamics of the transmembrane segment of phospholamban (TM-PLB), a sarcoplasmic Ca(2+)-regulator protein. (2)H NMR spectra of (2)H-labeled leucine (deuterated at one terminal methyl group) incorporated at different sites (CD(3)-Leu28, CD(3)-Leu39, and CD(3)-Leu51) along the TM-PLB peptide exhibited line shapes characteristic of either methyl group reorientation about the C(gamma)-C(delta) bond axis or by additional librational motion about the C(alpha)-C(beta) and C(beta)-C(gamma) bond axes. The (2)H NMR line shapes of all CD(3)-labeled leucines are very similar below 0 degrees C, indicating that all of the residues are located inside the lipid bilayer. At higher temperatures, all three labeled leucine residues undergo rapid reorientation about the C(alpha)-C(beta), C(beta)-C(gamma), and C(gamma)-C(delta) bond axes as indicated by (2)H line-shape simulations and reduced quadrupolar splittings. At all of the temperatures studied, the (2)H NMR spectra indicated that the Leu51 side chain has less motion than Leu39 or Leu28, which is attributed to its incorporation in the pentameric PLB leucine zipper motif. The (15)N powder spectra of Leu39 and Leu42 residues indicated no backbone motion, while Leu28 exhibited slight backbone motion. The chemical-shift anisotropy tensor values for (15)N-labeled Leu TM-PLB were sigma(11) = 50.5 ppm, sigma(22) = 80.5 ppm, and sigma(33) = 229 ppm within +/-3 ppm experimental error. The (15)N chemical-shift value from the mechanically aligned spectrum of (15)N-labeled Leu39 PLB in DOPC/DOPE phospholipid bilayers was 220 ppm and is characteristic of a TM peptide that is nearly parallel with the bilayer normal.     

Investigating structural changes in the lipid bilayer upon insertion of the transmembrane domain of the membrane-bound protein phospholamban utilizing 31P and 2H solid-state NMR spectroscopy

Phospholamban (PLB) is a 52-amino acid integral membrane protein that regulates the flow of Ca(2+) ions in cardiac muscle cells. In the present study, the transmembrane domain of PLB (24-52) was incorporated into phospholipid bilayers prepared from 1-palmitoyl-2-oleoyl-sn-glycero-phosphocholine (POPC). Solid-state (31)P and (2)H NMR experiments were carried out to study the behavior of POPC bilayers in the presence of the hydrophobic peptide PLB at temperatures ranging from 30 degrees C to 60 degrees C. The PLB peptide concentration varied from 0 mol % to 6 mol % with respect to POPC. Solid-state (31)P NMR spectroscopy is a valuable technique to study the different phases formed by phospholipid membranes. (31)P NMR results suggest that the transmembrane protein phospholamban is incorporated successfully into the bilayer and the effects are observed in the lipid lamellar phase. Simulations of the (31)P NMR spectra were carried out to reveal the formation of different vesicle sizes upon PLB insertion. The bilayer vesicles fragmented into smaller sizes by increasing the concentration of PLB with respect to POPC. Finally, molecular order parameters (S(CD)) were calculated by performing (2)H solid-state NMR studies on deuterated POPC (sn-1 chain) phospholipid bilayers when the PLB peptide was inserted into the membrane.     

Influence of protein conformation on its adaptability under chaotropic conditions

The thermostability of serum albumin and beta-lactoglobulin in various salt solutions was studied using differential scanning calorimetry. Below 1.0 M salt concentrations, the relative effectiveness of various sodium salts on increasing the thermostability of beta-lactoglobulin followed the classic Hofmeister or lyotropic series, i.e. SO2-(4) greater than Cl- greater than Br- greater than ClO-4 greater than SCN-; however, in the case of serum albumin the above order was reversed, i.e. ClO-4 greater than SCN- greater than Br- greater than Cl- greater than SO2-(4), indicating that the thermostability of serum albumin was higher in chaotropic solution conditions. Circular dichroic analysis of serum albumin in NaClO4 solutions revealed that the alpha-helical content of the protein increased from 59% to 73% in 1.0 M NaClO4; no similar increase in secondary structure was observed for beta-lactoglobulin. These observations contradicted the general notion that the chaotropic effect of neutral salts on the stability of macromolecules is independent of any details of the macromolecular conformation itself. The results presented here indicate that the predisposition of the native conformation of a protein per se might affect whether the protein would undergo stabilization or destabilization (i.e. conformational adaptability) under moderate chaotropic solution conditions.     

A comparison of DMPC- and DLPE-based lipid bilayers.

A 250 ps molecular dynamics simulation of the dimyristoylphosphatidylcholine (DMPC)-based lipid bilayer, including explicit water molecules, is reported. The solvent environment of the head groups and other structural properties of the bilayer have been analyzed and compared with experimental results as well as our previous simulation of the dilauroylphosphatidylethanolamine (DLPE)-based bilayer. From this comparison we find that the solvent structure around the DMPC head group (clathrate shell) is significantly different than that around the DLPE head group (typical hydrogen bonding interactions). We have modeled the probable relationship between the different solvent environments around the R-N(CH3)3+ (DMPC) and R-NH3+ (DLPE) head groups and the different interlammelar distances in these systems by performing potential of mean force (PMF) simulations on two N(CH3)4+ and NH4+ ions in water. From the PMF simulations it appears that the differences in the hydration of the DMPC and DLPE head groups is not responsible for the differences in the hydration force observed for these systems. We also find that the orientational polarization of DLPE and DMPC is similar, which suggests that solvent polarization is not responsible for the differences in the hydration repulsion behavior observed in these systems. We also examined the order parameters for DMPC and found them to be in reasonable agreement with experiment. Given the different characteristics of the DLPE and DMPC head groups, we suggest an explanation of the differences in the interlammellar spacings of bilayers composed of these like-charged lipids. From our DLPE simulations we find that the R-NH3+ head groups can interact with the nonesterified oxygens of the phosphate group in an intraleaflet or an interleaflet manner. For the latter a "cross link" between two leaflets can be formed, which causes a stabilization of the interlamellar spacings at fairly short distances. Moreover, due to the strong intraleaflet interaction we find that the DLPE interface is relatively "flat" (as opposed to DMPC-based bilayers), which results in a surface that has regions of positive and negative charge that reside in the same plane along the bilayer normal. Based on this we propose that the DLPE bilayer interface can correlate itself with another DLPE interface by alignment of the regions of positive (or negative) charge on one leaflet with the opposite charges on the opposing leaflet.     

Effect of alcohol feeding mode on the biosynthesis of poly (3-hydroxybutyrate-co-3-hyroxyvalerate)

An alcohol utilizing Alcaligenes eutrophus produced poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) copolymer under phosphate limitation. Fermentation was performed for 42-46 h in a computer-controlled 5-L working volume fed-batch fermentor using ethanol and propanol as carbon sources. The culture experienced phosphate limitation in approximately 19 h. When propanol was used as a sole carbon source, 24 g/L of copolymer with 36.5 mol % of hydroxyvalerate (HV) was produced at a polymer yield of 0.41 g polymer/g alcohol (g/g) and an average polymer production rate of 0.08 g polymer/g residual biomass-h (g/g-h). Two experiments switching alcohol after phosphate exhaustion resulted in better polymer production (g/L), polymer yield (g/g) on alcohol, HV yield (g/g) on propanol, and average polymer production rate (g/g-h) as compared to propanol run without alcohol switching. One switching experiment was from a mixture of 50% ethanol and 50% propanol to 100% propanol and the other experiment was from 100% ethanol to a mixture of 65% ethanol and 35% propanol. Polymer yield for these two experiments was 0.51 g/g and 0.46 g/g, respectively. However, HV mol % in the copolymer for these two runs (30.8 mol % 12.6 mol % respectively) was lower compared to propanol run without alcohol switching (3605 mol %). Direct switch from ethanol to propanol did not support cell growth and polymer production. Polymer production rate and polymer yield changed with time, and the pattern was dependent upon the alcohol feeding mode. (c) 1994 John Wiley & Sons, Inc.     

Solid-state nuclear magnetic resonance relaxation studies of the interaction mechanism of antimicrobial peptides with phospholipid bilayer membranes

An 18-residue peptide, KWGAKIKIGAKIKIGAKI-NH(2) was designed to form amphiphilic beta-sheet structures when bound to lipid bilayers. The peptide possesses high antimicrobial activity when compared to naturally occurring linear antimicrobial peptides, most of which adopt an amphipathic alpha-helical conformation upon binding to the lipids. The perturbation of the bilayer by the peptide was studied by static (31)P and (2)H solid-state NMR spectroscopy using POPC and POPG/POPC (3/1) bilayer membranes with sn-1 chain perdeuterated POPC and POPG as the isotopic labels. (31)P NMR powder spectra exhibited two components for POPG/POPC bilayers upon addition of the peptide but only a slight change in the line shape for POPC bilayers, indicating that the peptide selectively disrupted the membrane structure consisting of POPG lipids. (2)H NMR powder spectra indicated a reduction in the lipid chain order for POPC bilayers and no significant change in the ordering for POPG/POPC bilayers upon association of the peptide with the bilayers, suggesting that the peptide acts as a surface peptide in POPG/POPC bilayers. Relaxation rates are more sensitive to the motions of the membranes over a large range of time scales. Longer (31)P longitudinal relaxation times for both POPG and POPC in the presence of the peptide indicated a direct interaction between the peptide and the POPG/POPC bilayer membranes. (31)P longitudinal relaxation studies also suggested that the peptide prefers to interact with the POPG phospholipids. However, inversion-recovery (2)H NMR spectroscopic experiments demonstrated a change in the relaxation rate of the lipid acyl chains for both the POPC membranes and the POPG/POPC membranes upon interaction with the peptide. Transverse relaxation studies indicated an increase in the spectral density of the collective membrane motion caused by the interaction between the peptide and the POPG/POPC membrane. The experimental results demonstrate significant dynamic changes in the membrane in the presence of the antimicrobial peptide and support a carpet mechanism for the disruption of the membranes by the antimicrobial peptide.