前体、诱导子及抑制剂对紫杉烷生物合成的促进作用研究

研究了前体、诱导子和抑制剂对中国红豆杉细胞培养生产紫杉烷的促进作用。结果表明,向培养基中加入30mg/L 3-甲基－2－丁烯－1－醇,2mmol/L苯甲酸钠,10mg/L矮壮素,100μmol/L茉莉酸甲酯,0.1mmol/L丝氨酸,可以使紫杉醇含量增加1141.1%;加入0.1mmol/L苯甲酸钠,80μmol/L茉莉酸甲酯,0.1mmol/L丝氨酸可以使2α,5α,10β,14β－四乙酰氧基－紫杉－4（20）,11－二烯含量增加134.6%;25mg/L矮壮素,100μmol/L茉莉酸甲酯,0.5mmol/L丝氨酸可以使1β－羟基巴卡亭I含量增加95.2%;5mg/L3－甲基－2－丁烯－1－醇,10mg/L矮壮纱,40μmol/L茉莉酸甲酯,可以使14β-(2-甲基丁酰氧基）－2α,5α,10β－三乙酰氧基－紫杉－4（20）,11－二烯的含量增加76.4%。     

固定化微绿球藻去除NH4+-N、PO43——P效果的研究

为了探究固定化微绿球藻（Nannochloropsis oculata）去除污水中NH4+-N、PO43--P的效果,采用海藻酸钠固定化包埋技术进行实验。开展了固定化藻球大小、藻细胞包埋密度、藻球投放质量及充气培养条件对NH4+-N、PO43--P去除效果的单因子试验研究。结果表明,固定化藻球大小、藻细胞包埋密度、藻球投放质量和充气培养条件对NH4+-N、PO43--P的去除效果影响显著（P0.05）。藻球直径3.5 mm时生长速率（K）值最大（0.3320.002）,同时NH4+-N、PO43--P去除率效果最佳,分别为（75.083.83）%和（80.803.81）%;藻细胞包埋密度100104 cells/ball时K值最大（0.3300.033）,而NH4+-N、PO43--P去除率则以藻细胞包埋密度300104 cells/ball组为佳,分别达（87.200.43）%和（82.581.72）%,但考虑单位藻细胞去除率,包埋密度以100104 cells/ball为宜;随着藻球用量的增加K值下降,10 g/L组K值最大（0.3010.02）、50 g/L组K值最小（0.1930.01）,投放量30和50 g/L时NH4+-N去除率较高分别为（84.120.78）%和（84.630.45）%,30 g/L组PO43--P去除率最高达（77.131.43）%。综合考虑,藻球投放量选用30 g/L为宜;充气条件培养K值、NH4+-N和PO43--P去除率显著（P0.05）高于不充气,K值分别为（0.3060.006）和（0.1770.010）;NH4+-N去除率分别为（85.930.45）%和（49.320.45）%;PO43--P去除率分别为（66.665.00）%和（46.292.12）%。研究优化了微绿球藻固定化条件:固定化微绿球藻应进行充气培养,藻球规格3.5 mm、藻细胞包埋密度100104 cells/ball、藻球投放量30 g/L。     

EMS诱变和NaCl胁迫的正交实验及耐盐纤维亚麻种质的筛选

旨为经愈伤组织诱导再生的耐盐性纤维亚麻植株奠定基础。以3种纤维亚麻无菌苗的下胚轴段为材,经EMS诱变和盐胁迫的正交实验,再经诱芽和愈伤组织培养,研究其理化特性。结果显示,‘范妮’、‘天鑫3号’和‘双亚5号’下胚轴段不定芽培养的最适处理组合均为0.025%EMS处理2 h(或4 h),之后用100或150 mmol/L Na Cl处理12 h;总产芽率由高到低分别为‘范妮’、‘天鑫3号’和‘双亚5号’下胚轴段;对3种亚麻不定芽生长影响的顺序为:Na Cl浓度EMS浓度EMS处理时间;3种亚麻下胚轴段不定芽诱导的适宜培养基为MS+0.000 2 mg/L TDZ+0.02 mg/L KT+0.005 mg/L NAA+0.3%蔗糖+0.1 g/L肌醇+3.3 g/L植物凝胶。经0.025%EMS处理4 h+150 mmol/L Na Cl处理12 h后生长的愈伤组织,经过2次交替筛选后,其理化指标表明,与对照组的相比,3种亚麻处理组愈伤组织脯氨酸、可溶性糖、可溶性蛋白含量、SOD和POD活性均显著提高(P0.05)。经过0.025%EMS处理4 h+150 mmol/L Na Cl处理12 h后生长的愈伤组织具有一定的耐盐性。     

诱导子对百里香再生植株中苯丙氨酸解氨酶活性的影响

以1/2MS为基本培养基,分别用蔗糖、NO3-/NH4+、苯丙氨酸、谷氨酸和酪氨酸诱导培养百里香组培苗30d,以及茉莉酸甲酯(MeJA)、水杨酸(SA)和壳聚糖诱导培养百里香20、30和40d组培苗植株0～120h,在不同的时间点测定其苯丙氨酸解氨酶(PAL)活性,以探讨诱导子对增殖百里香中PAL活性的影响。结果显示:以1/2MS为基本培养基,碳氮源调控30d后,百里香组培株的PAL活性以蔗糖浓度为2%、NO3-/NH4+比例为2∶1时最高;前体调控添加0.6mmol/L苯丙氨酸、0.8mmol/L谷氨酸和0.4mmol/L酪氨酸处理的百里香组培苗的PAL活性最高,分别为相应对照的126.3%、150.6%和153.9%。培养20d和30d植株的PAL活性均以1.0mmol/L MeJA、100mg/L SA和200mg/L壳聚糖诱导处理的最高,分别为相应对照的610.3%、788.1%、592.5%以及402.4%、541.2%、400.1%;而40d植株经0.5mmol/L MeJA、100mg/L SA和200mg/L壳聚糖诱导的PAL活性最高,分别为对照的390.2%、377.1%和413.4%。可见,在不同时期添加不同的诱导子对百里香再生植株中PAL活性有显著影响,且不同材料和诱导子存在各自适宜的诱导浓度和最佳诱导时间。     

影响高山红景天细胞悬浮培养中细胞生长和红景天苷积累的几个因素(简报)

高山红景天细胞悬浮培养过程中,3mg·L916－BA、0．3mg·L－1NAA、60mmol·L－1氮源（其中NH_4~:NO_3~- 为1：1）、0．5～1．25mmol·L－1KH2PO4和200mg·L－1蛋白胨,较适合于细胞生长和红景天苷的积累。在适宜的接种量（3．2g·L-1）下细胞培养24d后,生物量达14．04g·L－1,干细胞中红景天苷含量为5．66mg·g-1。     

大豆磷酸烯醇式丙酮酸磷酸酯酶研究Ⅲ、非专一性酸性磷酸酯酶的纯化与特性

从大豆叶片中分离能水解磷酸烯醇式丙酮酸(PEP)的酸性磷酸酯酶,通过硫酸胺分部(20%～50%饱和度)沉淀、DEAE纤维素层析、刀豆球蛋白琼脂糖凝胶亲和层析将酶纯化了422.47倍,活性达78.16U/mg蛋白。该酶对PEP专一性不强,Km为1.09mmol/L(PEP),最适pH5.8,在pH4.8～7.0范围内及60℃以下较稳定,水解PEP的活性被Mg2+、Mn2+激活,F、Cu2+、Zn2+、PO43、MoO42及3磷酸甘油酸(3PGA)、三磷酸腺苷ATP等代谢物抑制,受异柠檬酸等有机酸影响较小。     

几种重金属离子对中华绒螯蟹（Eriocheir sinensis）N-乙酰-β-D-氨基葡萄糖苷酶活力的影响

本文研究了Cu2+、Pb2+、Zn2+和Ag+等重金属离子对中华绒螯蟹（Eriocheir sinensis）N-乙酰-β-D-氨基葡萄糖苷酶（NAGase）活力的影响。其结果表明：Cu2+、Pb2+和Zn2+对酶活力有不同程度的抑制作用, Cu2+和Zn2+对酶的抑制作用均表现为非竞争性抑制类型,Cu2+和Zn2+对酶的抑制常数（KI）分别为1.25 mmol/L和8.10 mmo/L;Pb2+对酶的抑制作用表现为混合型抑制类型,其对酶的抑制常数KI与KIS分别为10.44 mmol/L 和2.18 mmol/L。Ag+对酶的效应为先激活后抑制作用,其抑制作用表现为反竞争性抑制类型,Ag+对结合酶（ES）的抑制常数KI为204.51 mmol/L。     

重组枯草杆菌5-氨基酮戊酸脱水酶的纯化和性质研究

大肠杆菌中表达的枯草杆菌5-氨基酮戊酸脱水酶(5-aminolevulinate dehydratase, ALAD),其N端含有组氨酸标签,Ni-NTA 一步纯化至均一.纯化的酶比活为3.6 U/mg蛋白,酶的最适pH为8.0～10.0,Km为0.95 mmol,Vmax为16.7 mmol/h,在55 ℃温浴10 min,酶活保留85 %,2 mmol/L的K+、Zn2+、Mg2+、Li+、Fe3+和Mn2+提高酶活,2 mmol/L的Co2+和Ca2+对酶活没有显著影响,2 mmol/L的Cu2+和1 mmol/L的Hg2+完全抑制酶活.在2 mmol/L的EDTA存在下酶活性丧失,表明酶催化依赖金属离子.100 mmol/L的酮戊酸抑制酶活约50 %.10 mmol/L的2-巯基乙醇时提高酶活3倍,而100 mmol/L的二硫苏糖醇几乎完全抑制酶活.氨基酸修饰表明赖氨酸和半胱氨酸残基可能是酶催化必需的,组氨酸和丝氨酸残基对酶催化不起关键作用.     

反硝化除磷菌筛选及其特性研究

【目的】研究反硝化除磷菌特性。【方法】通过微生物筛选和生物学特性研究方法,从对虾养殖池塘中筛选出多株可在有氧条件下同时具有反硝化除磷功能的菌种。【结果】菌株LY-1可在18 h内将初始量为10 mg/L的亚硝酸盐氮降低至0.04 mg/L,PO43?-P降低至0.05 mg/L。在DO浓度为5.0?5.9 mg/L时,该菌反硝化除磷率近100%。试验选取具有反硝化除磷功能的枯草芽孢杆菌为阳性对照菌,大肠杆菌为阴性对照菌,比较研究了菌株LY-1在不同pH、温度、盐度、PO43?-P浓度、亚硝酸盐浓度时反硝化除磷的强弱,在pH为5?9范围时,该菌亚硝酸盐氮去除率近99%,PO43?-P去除率86%;温度为30°C时,该菌反硝化除磷率近100%;盐度为5‰?15‰、PO43?-P浓度为10 mg/L、亚硝酸盐氮浓度为20 mg/L时,该菌亚硝酸盐氮和PO43?-P去除率均可达99%。【结论】菌株LY-1反硝化除磷性能显著高于对照菌(P<0.05)。通过菌株LY-1形态学观察、生理生化及16S rRNA基因序列分析,初步鉴定为蜡样芽孢杆菌(Bacillus cereus)。     

三株野生大型真菌对孔雀石绿的脱色作用及其脱色条件

为了消除或降低孔雀石绿染料的污染,从50余种野生大型真菌中筛选到3种对孔雀石绿染料有较强脱色作用的菌株即落叶松附毛孔菌、粘小奥德蘑和皱盖囊皮菌。采用单因素和正交设计试验探索了优选菌株落叶松附毛孔菌的培养基组成和脱色条件。结果显示其最佳发酵培养基组成为马铃薯20%、玉米汁2%、KH2PO4 0.3%、MgSO4?7H2O 0.15%;最佳脱色条件为染料初始浓度100mg/L、发酵液用量6mL、Mg2+浓度6mmol/L、pH6.0、温度50℃、摇床振荡速度150r/min。此条件下对孔雀石绿染料脱色1h,脱色率可达92.6%,其中Fe2+对脱色有很强的抑制作用。发酵液脱色受热、pH和Fe2+影响,推测,落叶松附毛孔菌对孔雀石绿脱色的活性成分可能是漆酶。     

Determination of iodide using flow injection with acidic potassium permanganate chemiluminescence detection.

A simple and rapid flow-injection method is described for the determination of iodide, based on potassium permanganate chemiluminescence detection via oxidation of formaldehyde in aqueous hydrochloric acid. The calibration graph was linear over the range 1.0-12 x 10(-6) mol/L (r2 = 0.9955) with relative standard deviations (n = 4) in the range 1.0-3.5%. The detection limit (3sigma) was 1.0 x 10(-7) mol/L, with sample throughput of 120/h. The effect of interfering cations [Ca(II), Mg(II), Ni(II), Fe(II), Fe(III) and Pb(II)] and anions (Cl-, SO4(2-), PO4(3-), NO3-, NO2-, F- and SO3(2-)) were studied. The method was applied to iodized salt samples and the results obtained in the range 0.03 +/- 0.005 - 0.10 +/- 0.006 mg I/g were in reasonable agreement with the amount labelled. The method was statistically compared with the results obtained by titration; no significant disagreement at 95% confidence was observed.     

Anion inhibition of the proton pump in rat liver multivesicular bodies

Rat liver multivesicular bodies (MVB), as well as other hepatic subcellular organelles, are acidified by an electrogenic ATP-dependent proton pump that requires Cl- for maximal acidification (Van Dyke, R. W., Hornick, C. A., Belcher, J., Scharschmidt, B. F., and Havel, R.J. (1985) J. Biol. Chem. 260, 11021-11026), suggesting that Cl- serves as a permeable charge-compensating anion. However, we have observed that NO3- is unable to substitute for Cl-. This study was undertaken therefore to examine more closely the effects of Cl- on MVB acidification and to determine whether NO3- and other anions interact with the proton pump. ATP-dependent vesicle acidification and membrane potential (psi) were measured using the fluorescent dyes acridine orange and Oxonol V (bis(3-phenyl-5-oxoisoxasol-4-yl)pentamethine oxonol), respectively. Cl- both stimulated acidification (Km = 23.2 +/- 4.2 mM) and decreased psi (IC50 = 3.4 +/- 0.6 mM) in a concentration-dependent, nonlinear fashion. In the presence of saturating Cl- (100 mM), however, NO3- (shown to be more permeable than Cl-) and the impermeant anions SO4(2-) and PO4(2-), inhibited both ATP-dependent acidification and psi in a concentration-dependent manner. Other anions, including gluconate and HCO3-, had no effect. The inhibitory effect of NO3- was reversible. Neither SO4(2-) nor PO4(2-) appeared to block Cl- movement across the vesicle membrane as assessed by the ability of Cl- to decrease an established psi. In additional experiments, the effects of anions on relaxation of a previously established pH gradient were measured. Compared to Cl- or gluconate, NO3- had no significant effect on pH gradient relaxation, even when MVB were preloaded with NO3-, indicating that rapid cycling of NO3-/HNO3 across the MVB membrane does not occur. The organic nitrate, isosorbide dinitrate, also inhibited both acidification and psi and, similar to NO3-, had no effect on pH gradient relaxation. By contrast, NO2- potently inhibited both MVB acidification and psi but also rapidly relaxed a pre-established pH gradient, suggesting that NO2- increases MVB membrane proton permeability. Finally, MVB exhibited N-ethylmaleimide-sensitive ATPase activity that was inhibited 23.9% by NO3- (100 mM). In conclusion, although MVB are permeable to a variety of anions (Cl-, Br-, NO3-, NO2-), only Cl- and Br- support maximal rates of acidification by the proton pump.(ABSTRACT TRUNCATED AT 400 WORDS)     

Ionic regulation of glutamate binding sites

Cl- and Ca2+ increase glutamate binding to rat synaptic plasma membranes (SPMs) by revealing a distinct class of L-glutamate (L-Glu) binding sites. The present study was conducted to examine both the anion specificity of this response and the nature of the interaction between Cl- and Ca2+. Of the anions tested, Br- was the most effective in increasing the levels of L-Glu binding. Other effective anions were Cl-, NO3- and formate while F-, HCO3-CIO4-, propionate, SO42- and PO43- were ineffective. The anion specificity was similar to that observed for the Cl- membrane channel, suggesting that this binding site and the ion channel may be related. In the absence of Cl-, Ca2+ has little effect on L-Glu binding. Increasing the Cl- concentration increased the apparent affinity (decreased KCa2+) of the Ca2+-stimulated, L-Glu binding component and also increased the maximal amount of the enhancement. Conversely, increasing Ca2+ levels increased the maximal enhancement of L-Glu binding brought about by Cl- without affecting the KCl- of the effect. Prior incubation of membranes with Ca2+ did not raise the level of L-Glu binding. Furthermore, EGTA was able to reverse the stimulation of L-Glu binding due to Ca2+. The results indicate that Ca2+ acts ionically to enhance L-Glu binding to rat SPMs.     

Anion recognition by thiostrepton

A bicyclic polypeptide antibiotic thiostrepton forms both 1:1 and 1:2 complexes with anions (as tetrabutylammonium salts) in organic solvents with K2 >K1 for F- and K2Cl-, Br-, HSO4-, H2PO4-, but in CHCl3 they follow a different order: Cl- approximatelyHSO4- >F- approximately AcO- > Br > H2PO4-. The binding mode of anions to thiostrepton is discussed on the basis of solvent effects on the complexation selectivity.     

The influence of inorganic anions on the formation and stability of Fe3+-transferrin-anion complexes

Harris (Biochemistry 24 (1985) 7412) reports that inorganic anions bind to human apotransferrin in such a way as to perturb the ultraviolet spectrum. The locus of binding is thought to involve the specific metal/anion-binding sites since no perturbation is observed with Fe3+-transferrin-CO3(2-). Paradoxically, we were unable to demonstrate the formation of Fe3+-transferrin-inorganic anion complexes despite the presence of high concentrations of SO4(2-), H2PO4-, Cl-, ClO4- or NO3-. Similar results were found for human lactoferrin. Electron paramagnetic resonance spectroscopy and visible spectrophotometry were used to monitor the results. An attempt to form the H2PO4- complex by displacement of glycine from Fe3+-transferrin-glycine resulted only in the disruption of the ternary complex. A series of inorganic anions varied in their ability to release iron from Fe3+-transferrin-CO3(2-) at pH 5.5, the approximate pH of endosomes where iron release takes place within cells. The order of effectiveness was H2P2O7(2-) much greater than H2PO4- greater than SO4(2-) greater than NO3- greater than Cl- greater than ClO4-. The rate of iron removal from Fe3+-transferrin-CO3(2-) at pH 5.5 by a 4-fold excess of pyrophosphate was greatly enhanced by physiological NaCl concentration. Iron removal was complete within 10 min, the approximate time for iron release from Fe3+-transferrin-CO3(2-) in developing erythroid cells. Thus, inorganic anions may have a significant effect on the release of iron under physiological conditions despite the fact that such inorganic anions cannot act as synergistic anions. The results are discussed in relation to a special role for the carboxylate group in allowing ternary complex formation.     

Influence of nitrate and COD on phosphorus, nitrogen and dinitrotoluene (DNT) removals under batch anaerobic and anoxic conditions

In this study, the effects of COD to NO(3)-N ratio in the feed on PO(4)-P removal was investigated. Maximum PO(4)-P uptake was obtained in the anoxic reactor when the COD to NO(3)-N ratios were between 2 and 3.75. With the influent COD of 800-1500 mg COD/L a total of the maximum removable PO(4)-P was 56 mg PO(4)-P/L through 20 days of anaerobic/anoxic incubation, indicating 98% P removal in the anoxic reactor. Similarly, for the COD to NO(3)-N ratios varying between 2 and 3.75 maximum denitrification was observed. Through anoxic operation the poly-P bacteria are capable of removing NO(3)-N using VFA, COD as carbon source and NO(3)-N as the electron acceptor after methanogenesis has been completed. High NO(3)-N concentrations stopped significantly the P uptake. A total of 97-99% dinitrotoluene removal efficiencies in the reactors containing COD to NO(3)-N ratio of 2 and 3.75 after 20 days of incubation period. For maximum NO(3)-N and PO(4)-P removals optimal COD to NO(3)-N ratios, COD and NO(3)-N concentrations were 2-3.75, 2000-4000 mg COD/L and, 800-1500 mg NO(3)-N/L, respectively.     

Effects of extracellular anions on cell growth of Chinese hamster V-79 cells

The effects of extracellular anions (10-150 mM, added as Na salts to normal growth medium) on the growth of Chinese hamster V-79 cells were examined. Additions of NaCl and NaNO3 at concentrations greater than 60 mM reduced the growth rate dose-dependently. Several other anions also inhibited cell growth in the decreasing order of potency, SCN- greater than NO2- greater than NO3- greater than Br- greater than Cl- greater than gluconate- glutamate- greater than Mes-. When the added anions were removed, the growth rate was restored to the control rate. Cell survival was markedly reduced by the addition of SCN-, but was less affected by other anions (Cl-,NO3- and NO2-) of comparable potency. The respective syntheses of cellular DNA and protein, as estimated from the incorporation of [3H]-thymidine and [14C]leucine, also decreased with the increase in the concentration (60-120 mM) of anions added, the order of potency being SCN- greater than NO2- greater than NO3- greater than Cl-. After anion-treatment, the cellular Na+ concentration increased and the cellular Cl- concentration decreased in the order of SCN- greater than NO2- greater than NO3-, Cl-, but, the cellular K+ concentration did not change significantly. These data suggest that changes in extracellular anions affect cell growth and survival, probably through changes in the intracellular Na+ or Cl- concentration and in the rates of protein and/or DNA synthesis.     

Determination of iron in blood serum using flow injection with luminol chemiluminescence detection.

A simple and rapid fl ow injection method is reported for the determination of iron in blood serum after acid digestion with HNO3 and HClO4, based on luminol CL detection in the absence of added oxidant. The detection limit (3 s) was 1.0 nmol/L with a sample throughput of 120/h. The calibration graph was linear over the range 0.001-1.0 micromol/L (r2 = 0.9974), with relative standard deviations (RSD) (n = 4) in the range 3.2-5%. The effect of interfering cations (Ca(II), Mg(II), Cu(II), Cd(II), Pb(II), Mn(II), Zn(II), Ni(II), Co(II) and Fe(III)) and anions (Cl-, SO4(2-), HCO3-, NO3-, NO2-) were studied using a luminol CL system for Fe(II) determination. The method was applied to normal blood serum and the results (1.32 +/- 0.08-1.74 +/- 0.05 mg/L) were compared with those from a spectrophotometric reference method (1.34 +/- 0.06-1.80 +/- 0.10 mg/L), which agree fairly well with the overall reference range in blood.     

Lysis of Streptococcus mutans by hen egg white lysozyme and inorganic sodium salts.

Streptococcus mutans BHT was grown in a synthetic medium containing radioactive thymidine to monitor deoxyribonucleic acid release. Kinetic experiments demonstrated that although lysozyme alone could not liberate deoxyribonucleic acid, cellular deoxyribonucleic acid was liberated from lysozyme-treated cells by addition of low concentrations of inorganic sodium salts. When the salts were tested for their ability to dislodge cell-bound tritiated lysozyme, the extent of the initial release of enzyme by individual anions correlated with the anion potency for deoxyribonucleic acid liberation (SCN- greater than ClO4- greater than I- greater than Br- greater than NO3- greater than Cl- greater than F-), although the total amount of lysozyme dislodged did not correspond directly with cell lysis. Differences in the effectiveness of anions (SCN-, HCO3-, Cl- and F-) in potentiating cell lysis could be enhanced or minimized by varying the lysozyme, anion, and bacterial cell concentrations. As the anion concentration was increased for each enzyme concentration and cell concentration, the lysis increased, in some cases markedly, until maximum levels of released deoxyribonucleic acid were attained. The maximum levels of lysis of SCN- and HCO3- were similar and were greater than those for Cl- and F-. In addition, the maximum levels were observed to increase for each of the anions as the concentration of lysozyme increased.     

The effect of Cl- depletion and X- reconstitution on the oxygen-evolution rate, the yield of the multiline manganese EPR signal and EPR signal II in the isolated Photosystem-II complex

The role of Cl- in photosynthetic O2 evolution has been investigated by measurement of the steady-state O2 rate and EPR of the electron donors responsible for the S2 multiline signal and Signal IIs upon Cl- depletion and substitution in Photosystem II membranes. Cl- removal has three effects upon the donor side of Photosystem II. (1) It abolishes O2 evolution reversibly, while decreasing the yield of the S2 multiline signal indicative of the manganese site of the O2-evolving complex in the S2 oxidation state. This decrease is brought about by (2) the reversible disconnection of the manganese complex from the reaction center; and by (3) deactivation of S1 centers having reduced primary acceptor QA to form SO centers having a reduced Signal IIs species. Reactivation of O2 evolution by anions confirms earlier work showing a requirement for a univalent anion of optimum charge density. The observed order of reactivation is Cl- greater than Br- approximately NO3- much greater than OH- approximately F-. Reactivation of the S2 multiline signal follows Cl- approximately Br- greater than NO3- approximately OH- greater than F-, in near correspondence with reactivation of O2-evolution rates. Cl- titrations of F- -inhibited samples reveal two binding sites for Cl- which differ in binding affinity by 11-fold. The higher-affinity site reactivates the S1----S2 light reaction, while the lower-affinity site reactivates the S3----S0 light reaction. The high affinity site is located within the O2-evolving complex at an undetermined site, while the lower-affinity site functions in coupling the reaction center photochemistry to the O2-evolving complex. The results are compared with Cl-/F- exchange equilibria for Mn3+ in solution. A model for the lower S-state transitions is presented in which specific oxidation state assignments are made for some of the donors and acceptors of Photosystem II.