降水与CO2浓度协同作用对短花针茅光合特性的影响

降水变化与CO2浓度升高将严重影响陆地生态系统尤其是草地生态系统,阐明干旱半干旱区草原优势植物对降水与CO2浓度变化的联合响应有助于理解和准确评估未来气候变化对草地生态系统的影响.基于开顶式生长箱(OTC),模拟研究了降水变化(-30％、-15％、0、+15％、+30％(以1978-2007年月降水平均值为基准))、CO2浓度变化(对照、450 μmol·mol-1、550 μmol·mol-1)及其协同作用对荒漠草原优势物种短花针茅(Stipa breviflora)光合特性的影响.结果表明:降水变化和CO2浓度升高对短花针茅光合参数影响显著,表现出显著的交互作用.随着CO2浓度升高,短花针茅叶片净光合速率(Pn)呈增加趋势,但随着时问延长(8月份)显示出光合适应现象;气孔导度(Gs)和蒸腾速率(Tr)则呈下降趋势,水分利用效率(WUE)显著增加.随着降水增加,短花针茅的Pn、Gs和Tr均呈增加趋势,Pn增加速率小于Tr,使得WUE降低.高浓度CO2和降水增加15％的协同作用可以显著提高短花针茅的Pn、Gs和Tr,但Pn增加速率接近于Tr,导致WUE变化不显著.这表明,在干旱半干旱地区,CO2浓度升高可在一定程度上提高短花针茅的抗旱能力,增强短花针茅对暖干化气候情景的适应性.     

内蒙古地带性针茅植物对CO2和气候变化的适应性研究进展

收集了1992—2013年关于模拟CO2浓度升高及气候变化(温度升高、降水变化)对内蒙古地带性草原群落的5个建群种针茅植物(贝加尔针茅、本氏针茅、大针茅、克氏针茅、短花针茅)影响的实验研究结果表明,模拟CO2浓度升高、增温和增雨将提高针茅植物的光合作用和株高生长,但CO2处理时间延长会导致光合适应;温度和降雨变化将改变针茅植物的物候进程,但物种之间反应有差异;CO2浓度升高有助于针茅植物生物量增加,增温和干旱则相反,CO2浓度升高对干旱的影响具有补偿作用;干旱和涝渍胁迫将提高针茅植物植株C/N,CO2浓度升高将加剧水分胁迫下针茅植物植株C/N的增加效应,导致牧草品质下降。由于当前在适应性指标、针茅植物对气候变化协同作用的适应机理及其敏感性研究等方面存在的不足,导致目前无法全面比较各针茅植物对CO2和温度、降水变化的响应差异及其敏感性,因而无法预测未来在全球变化背景下,这几种针茅植物的动态变化及其在地理分布上的迁移替代规律。为科学应对气候变化,未来应加强内蒙古地带性针茅植物的适应性指标、针茅植物对多因子协同作用的适应机理及敏感性研究。     

温度与降水协同作用对短花针茅生物量及其分配的影响

采用人工气候箱法模拟研究温度变化(对照、增温1.5、2.0、4.0、6.0℃)、降水变化(W-30%、W-15%、对照W0、W+15%、W+30%)(以1978—2007年6—8月的月均温和月均降水量为对照)及其协同作用对内蒙古荒漠草原优势种短花针茅生物量及其分配的影响表明:温度和降水协同作用显著影响短花针茅生物量及其分配。增温使短花针茅总生物量、叶和根生物量增加,茎生物量和根冠比分别在在增温1.5℃、4℃后随增温而减小。干旱高温使总生物量减小,增温和降水增加使总生物量、根和叶生物量增加,茎生物量在增温小于2℃时随降水增加而增加,则适当的增温和增加降水可促进短花针茅生物量的积累。短花针茅的各器官生物量分配对温度和降水协同作用的敏感性为茎生物量比叶生物量比根生物量比,最敏感性器官为茎。这表明,短花针茅可根据不同器官生物量对水热变化的敏感性,调节干物质累积分配来适应气候变化。     

内蒙古不同草地类型针茅属植物对放牧强度和气候因子的响应

以内蒙古草甸草原、典型草原、荒漠草原为研究对象,设置轻度、中度、重度以及对照4个放牧梯度,经过连续5年野外调查试验,研究针茅属植物在不同放牧强度及不同水热条件下的变化趋势,为针茅属植物为建群种的草地管理提供理论基础。结果表明:1)草甸草原的贝加尔针茅相对生物量随着放牧强度的增加呈下降趋势,即使降水波动较大,依然保持该趋势;2)典型草原大针茅受降水的影响较大,在对照样地中相对生物量与降水呈显著正相关(r=0.326),在轻度放牧样地中则为负相关(r=-0.319),在干旱年份大针茅相对生物量在轻度放牧强度中显著高于对照样地,所以放牧对大针茅的影响随降水的不同而不同;3)荒漠草原的短花针茅相对生物量随放牧强度增加而降低,但差异不显著,短花针茅受降水和温度的影响较大,与降水呈显著负相关(r=-0.440~-0.583),与温度呈显著正相关(r=0.282~0.299)。综上所述,在水分不受限制的草甸草原,即使降水温度有较大波动也不会对群落建群种产生较大影响,放牧成为草甸草原退化的主要因子;而在降水较少的典型草原和荒漠草原,气候逐渐成为除放牧以外的主要制约因子。     

不同载畜率下短花针茅分蘖数量对内源激素浓度的响应

放牧是荒漠草原最主要的利用方式之一,载畜率的变化严重影响着植物的生长发育;而内源植物激素是调节植物生长发育的开关,且植物在不同的生长发育阶段具有不同的生理要求和环境适应能力。通过测定放牧条件下短花针茅(Stipa breviflora)分蘖叶内源激素的变化,研究短花针茅分蘖生长对放牧的响应,并分析了分蘖数量受内源激素影响的机制。结果表明,(1)内源激素与载畜率之间存在显著二次相关关系,说明放牧能够显著增加内源激素的浓度(P0.05),但这种相关只存在于中、小株丛的短花针茅中。(2)其次,放牧能够在一定程度上影响短花针茅植株个体分蘖的数量(P0.05),重度放牧是增加短花针茅植株个体分蘖数量最显著的载畜率。(3)过高浓度的生长素(IAA)会抑制短花针茅的分蘖数量(P0.01)。而细胞分裂素(CTK)与短花针茅的分蘖数量之间尚未发现相关关系。     

长期降水量变化下荒漠草原植物生物量、多样性的变化及其影响因素

植物多样性是植物群落维持生态系统稳定的基础。虽然荒漠草原植被稀少,但其在防风固沙等方面仍发挥着不可替代的生态服务功能,然而对荒漠植物多样性如何响应长期极端降水量变化尚缺乏深入理解。该研究依托2014年在宁夏荒漠草原设立的降水量变化(减少50%、减少30%、自然、增加30%和增加50%)的野外实验样地,研究了2020年5–10月植物生物量和物种多样性的变化特征,分析了二者与土壤性质的关系。随着生长季推移,植物群落生物量、Patrick丰富度指数和Shannon-Wiener多样性指数呈先增加后降低的时间动态,Pielou均匀度指数和Simpson优势度指数无明显的变化规律。与自然降水量相比,降水量减少对植物生物量和多样性影响较小,尤其是降水量减少30%的处理下;多数情况下,降水量增加刺激了苦豆子(Sophora alopecuroides)、短花针茅(Stipa breviflora)、白草(Pennisetum centrasiaticum)等物种生长,提高了植物生物量,但亦未明显改变植物多样性(尤其是降水量增加30%的处理下)。对植物生物量影响显著的土壤因子包括脲酶活性、温度、含水量...     

CO2浓度升高和降水增加协同作用对玉米产量及生长发育的影响

关于[CO₂]升高和降水变化等多因子共同作用对植物的影响报道较少, 制约着人们对植物对全球气候变化响应的认识和预测。玉米(Zea mays)作为重要的C₄植物, 受[CO₂]和降水影响显著, 但鲜有[CO₂]升高和降水增加协同作用对其产量及生长发育影响的报道。该研究利用开顶式生长箱模拟[CO₂]升高(390 (环境)、450和550 μmol·mol^-1), 降水增加量设置为增加自然降水量的15% (以试验地锦州1981-2010年6至8月月平均降水量为基准), 从而形成6个处理: C₅₅₀W_+15%、C₅₅₀W₀、C₄₅₀W_+15%、C₄₅₀W₀、C₃₉₀W_+15%和C₃₉₀W₀。试验材料选用玉米品种‘丹玉39’。结果表明: [CO₂]升高和降水增加的协同作用在玉米的籽粒产量和生物产量上均达到了显著水平(p< 0.05), 二因子均起正作用, 使籽粒产量和生物产量均升高。籽粒产量在[CO₂] 390、450和550 μmol·mol^-1水平下的降水增加处理较自然降水处理分别增加15.94%、9.95%和9.45%, 而生物产量分别增加13.06%、8.13%和6.49%。因为籽粒产量的增幅略大于生物产量的增幅, 所以促进了经济系数的升高。穗部性状变化显著, 其中, 穗粒数、穗粒重、穗长和穗粗等性状值均随[CO₂]升高而升高, 且各[CO₂]水平下均表现为降水增加处理>自然降水处理, 而瘪粒数相反。但是, [CO₂]升高和降水增加的协同作用也促进了轴粗的升高, 对玉米产量的增加起着限制作用。二因子协同作用在净光合速率(P_n)和叶面积上达到了极显著水平(p< 0.01), 而在株高和干物质积累量上达到了显著水平(p< 0.05)。二因子协同作用使玉米叶片的P_n升高, 植株高度升高, 穗位高升高, 茎粗增加, 叶面积变大, 从而促进了干物质积累量的升高, 为玉米增产打下了良好的基础。这表明: 在未来[CO₂]升高条件下, 一定程度的降水增加对玉米的产量具有正向促进作用。     

CO2浓度升高和不同灌溉量对东北玉米光合特性及产量的影响

CO2和水分是植物光合作用的重要底物,大气CO2浓度升高或水分变化影响植物光合作用。玉米是重要的C4植物,目前已成为我国第一大作物。我国东北地区的玉米产量占全国玉米产量的1/3左右,对确保国家的粮食安全具有重要作用。但是,关于CO2浓度升高或水分变化共同作用对东北玉米的光合速率、水分利用效率和产量影响的研究甚少。基于开顶式生长箱(OTCs),模拟研究了CO2浓度变化(390、450、550μmol/mol)和降水变化(0、+15%(以试验地锦州1981—2010年6、7、8月月平均降水量88.7,153.9 mm和139.8 mm为基准))共同作用对玉米光合特性及产量的影响。以玉米品种丹玉39为材料,利用直角双曲线修正模型对6个处理(C550W+15%、C550W0、C450W+15%、C450W0、C390W+15%和C390W0)的光响应曲线进行了拟合。结果表明:在CO2浓度升高和灌溉的共同作用下,玉米叶片净光合速率(Pn)升高,且灌溉作用大于高CO2浓度作用;而蒸腾速率(Tr)则下降,使水分利用效率(WUE)升高。CO2浓度升高使气孔导度(Gs)降低,灌溉则使之升高,但灌溉的作用小于高CO2浓度作用;胞间CO2浓度(Ci)随CO2浓度增加而升高,灌溉对其影响不明显。高CO2浓度和灌溉共同作用下光响应参数差异明显。CO2浓度升高增加了最大净光合速率(Pnmax)和光饱和点(LSP),灌溉亦然;CO2浓度升高使得光补偿点(LCP)、光补偿点量子效率(φc)和暗呼吸速率(Rd)的灌溉处理和自然降水处理的差距变小。390、450、550μmol/mol CO2浓度下的灌溉处理与自然降水处理相比,叶面积分别增加了11.56%、3.31%和0.45%,干物质积累量分别增加了14.69%、8.09%和1.01%,最终使产量分别增加了10.47%、12.07%和8.96%。可见,在高CO2浓度下,适量的灌溉对玉米的整个光合作用过程起到了促进作用,最终表现为籽粒产量的增加。为研究者评估气候变化对中国东北地区作物光合能力和产量的影响及决策者调整适应气候变化措施方面提供依据。     

荒漠草原建群种短花针茅功能性状对长期放牧的可塑性响应

植物功能性状反映了植物对生长环境的响应和适应,将环境、植物个体和生态系统结构、过程与功能联系起来,而放牧是草原生态系统的主要土地利用方式,对草原生态系统的群落结构和生态系统功能的改变起着重要作用。该研究测定了内蒙古四子王旗长期不同放牧强度下草原建群种短花针茅(Stipa breviflora)的功能性状,探讨放牧对草原群落结构的影响。研究发现:(1)随放牧强度增加,叶片各性状指标、个体生物量、植株高度、冠(丛)幅均有显著变小、变矮的趋势。(2)通过排序构建的短花针茅性状可塑性变化谱显示,各功能性状指标的可塑性指数均在重牧区最高,轻牧区最低;茎质量、叶质量、全株地上生物量对放牧响应敏感,而茎基部直径、冠(丛)幅和比叶质量响应不敏感。(3)短花针茅对长期放牧的变异系数(CV)与其响应程度(PI)之间符合指数方程y=0.08+0.18e~(1.61 x)(R~2=0.53,P0.05),植物功能性状的变异性与性状对放牧的响应强度呈正相关关系。(4)影响地上生物量的因子中,表型功能性状中的冠(丛)幅、平均叶长、叶片数量和总叶面积的贡献率之和达到72.93%。研究认为,随着放牧强度增强,短花针茅的变异性增大、可塑性增强,这可能是短花针茅能够成为建群种的生态响应机制。     

控制降水梯度对荒漠草原优势植物叶功能性状及土壤养分的影响

探究植物叶功能性状随降水梯度的变化规律,对揭示干旱区优势植物对环境变化的响应和适应策略至关重要。以盐池荒漠草原为研究对象,采用遮雨棚和喷灌系统控制降水梯度,分析了优势植物蒙古冰草（Agropyron mongolicum）、短花针茅（Stipa breviflora）及达乌里胡枝子（Lespedeza davurica）叶功能性状变异,以及土壤水分、养分和微生物特性与响应性状间的相关关系。结果表明：HW（增水50%）处理下3个植物LA（叶面积）、LDMC（干物质含量）显著增大,LW处理（减水50%）下短花针茅和达乌里胡枝子LA、LDMC显著减小,降水处理对短花针茅和达乌里胡枝子SLA（比叶面积）影响不显著,LW处理显著提高了蒙古冰草SLA;LW处理显著提高了蒙古冰草和短花针茅LNC（叶氮含量）和LPC（叶磷含量）;HW显著降低了土壤C、N含量,LW和HW均显著减少了真菌数量,而放线菌数量、微生物生物量C、N显著增加;3种优势植物LA均与土壤水分显著正相关,蒙古冰草和短花针茅通过提高SLA、LNC及LNP来适应干旱生境,蒙古冰草和短花针茅LNC及LNP是表征土壤P、微生物生物量有效性的关键指标,达乌里胡枝子通过自我调节养分利用策略来适应C、N、P含量和微生物活性较低的生境,从而决定其在群落中的优势地位。     

Tuning of functional heme reduction potentials in Shewanella fumarate reductases

The fumarate reductases from S. frigidimarina NCIMB400 and S. oneidensis MR-1 are soluble and monomeric enzymes located in the periplasm of these bacteria. These proteins display two redox active domains, one containing four c-type hemes and another containing FAD at the catalytic site. This arrangement of single-electron redox co-factors leading to multiple-electron active sites is widespread in respiratory enzymes. To investigate the properties that allow a chain of single-electron co-factors to sustain the activity of a multi-electron catalytic site, redox titrations followed by NMR and visible spectroscopies were applied to determine the microscopic thermodynamic parameters of the hemes. The results show that the redox behaviour of these fumarate reductases is similar and dominated by a strong interaction between hemes II and III. This interaction facilitates a sequential transfer of two electrons from the heme domain to FAD via heme IV.     

Electron transfer kinetics between soluble modules of Paracoccus denitrificans cytochrome c1 and its physiological redox partners

The transient electron transfer (ET) interactions between cytochrome c₁ of the bc₁-complex from Paracoccus denitrificans and its physiological redox partners cytochrome c₅₅₂ and cytochrome c₅₅₀ have been characterized functionally by stopped-flow spectroscopy. Two different soluble fragments of cytochrome c₁ were generated and used together with a soluble cytochrome c₅₅₂ module as a model system for interprotein ET reactions. Both c₁ fragments lack the membrane anchor; the c₁ core fragment (c_1CF) consists of only the hydrophilic heme-carrying domain, whereas the c₁ acidic fragment (c_1AF) additionally contains the acidic domain unique to P. denitrificans. In order to determine the ionic strength dependencies of the ET rate constants, an optimized stopped-flow protocol was developed to overcome problems of spectral overlap, heme autoxidation and the prevalent non-pseudo first order conditions. Cytochrome c₁ reveals fast bimolecular rate constants (10⁷ to 10⁸ M^− 1 s^− 1) for the ET reaction with its physiological substrates c₅₅₂ and c₅₅₀, thus approaching the limit of a diffusion-controlled process, with 2 to 3 effective charges of opposite sign contributing to these interactions. No direct involvement of the N-terminal acidic c₁-domain in electrostatically attracting its substrates could be detected. However, a slight preference for cytochrome c₅₅₀ over c₅₅₂ reacting with cyochrome c₁ was found and attributed to the different functions of both cytochromes in the respiratory chain of P. denitrificans.     

Thermodynamic and kinetic characterisation of individual haems in multicentre cytochromes c3

The characterisation of individual centres in multihaem proteins is difficult due to the similarities in the redox and spectroscopic properties of the centres. NMR has been used successfully to distinguish redox centres and allow the determination of the microscopic thermodynamic parameters in several multihaem cytochromes c₃ isolated from different sulphate-reducing bacteria. In this article we show that it is also possible to discriminate the kinetic properties of individual centres in multihaem proteins, if the complete microscopic thermodynamic characterisation is available and the system displays fast intramolecular equilibration in the time scale of the kinetic experiment. The deconvolution of the kinetic traces using a model of thermodynamic control provides a reference rate constant for each haem that does not depend on driving force and can be related to structural factors. The thermodynamic characterisation of three tetrahaem cytochromes and their kinetics of reduction by sodium dithionite are reported in this paper. Thermodynamic and kinetic data were fitted simultaneously to a model to obtain microscopic reduction potentials, haem-haem and haem-proton interacting potentials, and reference rate constants for the haems. The kinetic information obtained for these cytochromes and recently published data for other multihaem cytochromes is discussed with respect to the structural factors that determine the reference rates. The accessibility for the reducing agent seems to play an important role in controlling the kinetic rates, although is clearly not the only factor.     

The Rieske/cytochrome b complex of Heliobacteria

Heliobacteria have a Rieske/cytochrome b complex composed of a Rieske protein, a cytochrome b_6, a subunit IV and a di-heme cytochrome c. The overall structure of the complex seems close to the b₆f complex from cyanobacteria and chloroplasts to the exception of the di-heme cytochrome. We show here by biochemical and biophysical studies that a heme c_i is covalently attached to the Rieske/cytochrome b complex from Heliobacteria. We studied the EPR signature of this heme in two different species, Heliobacterium modesticaldum and Heliobacillus mobilis. In contrast to the case of b₆f complex, a strong axial ligand to the heme is present, most probably a protonatable amino acid residue.     

Spatio-temporal variability in the photosynthetic characteristics of Zostera tasmanica measured by PAM

Rapid light curves (RLCs), based on pulse amplitude modulated (PAM) fluorometry, were used to investigate the spatio-temporal variability in photosynthesis versus irradiance parameters (α, I_k and P_max) and the F_v/F_m ratio of the seagrass Zostera tasmanica (formerly Heterozostera tasmanica). Spatial variation was examined across scales ranging from within a leaf (cms) to across the bed (ms), using a nested analysis of covariance sampling design. Overall, significant variation was identified at all scales examined, excluding the largest scale (area). Patterns of variability differed among individual parameters; however a high percentage of the variation was consistently assigned to the covariates, age (within and between leaves) for all parameters, except P_max.     

Functional properties of type I and type II cytochromes c3 from Desulfovibrio africanus

Type I cytochrome c₃ is a key protein in the bioenergetic metabolism of Desulfovibrio spp., mediating electron transfer between periplasmic hydrogenase and multihaem cytochromes associated with membrane bound complexes, such as type II cytochrome c₃. This work presents the NMR assignment of the haem substituents in type I cytochrome c₃ isolated from Desulfovibrio africanus and the thermodynamic and kinetic characterisation of type I and type II cytochromes c₃ belonging to the same organism. It is shown that the redox properties of the two proteins allow electrons to be transferred between them in the physiologically relevant direction with the release of energised protons close to the membrane where they can be used by the ATP synthase.     

Generation, characterization and crystallization of a highly active and stable cytochrome bc1 complex mutant from Rhodobacter sphaeroides

The availability of the three dimensional structure of mitochondrial enzyme, obtained by X-ray crystallography, allowed a significant progress in the understanding of the structure-function relation of the cytochrome bc₁ complex. Most of the structural information obtained has been confirmed by molecular genetic studies of the bacterial complex. Despite its small size and simple subunit composition, high quality crystals of the bacterial complex have been difficult to obtain and so far, only low resolution structural data has been reported. The low quality crystal observed is likely associated in part with the low activity and stability of the purified complex. To mitigate this problem, we recently engineered a mutant [S287R(cytb)/V135S(ISP)] from Rhodobacter sphaeroides to produce a highly active and more stable cytochrome bc₁ complex. The purified mutant complex shows a 40% increase in electron transfer activity as compared to that of the wild type enzyme. Differential scanning calorimetric study shows that the mutant is more stable than the wild type complex as indicated by a 4.3 °C increase in the thermo-denaturation temperature. Crystals formed from this mutant complex, in the presence of stigmatellin, diffract X-rays up to 2.9 Å resolution.     

异育银鲫GABAB受体1亚基cDNA部分序列的克隆及表达分析

为了确定γ-氨基丁酸B受体（gamma-aminobutyric acid B receptor,GABA_BR）基因在异育银鲫（Carassius auratus gibelio）不同组织中的表达,本实验分别对异育银鲫不同组织中GABA_BR1基因进行RT-PCR扩增,并进行了克隆和测序,在与GenBank基因库中已知GABA_BR1序列进行同源性比对的基础上采用邻接法构建系统发育树,并进一步分析其在异育银鲫不同组织内的表达水平。经克隆获得异育银鲫GABA_BR1基因CDS区序列383 bp,编码127个氨基酸。荧光定量PCR结果显示,GABA_BR1基因在异育银鲫脑、肝、肾、心、肠、鳔、鳃、肌、尾鳍、脾、卵巢、精巢组织中均有表达,且在不同组织中的表达水平由高到低依次是：脑>尾鳍>精巢>心、肠、鳔>卵巢、脾、鳃、肌>肝、肾。本研究证实了GABA_BR1基因在异育银鲫各组织中表达的广泛性,且有明显的组织特异性。     

Evidence of functional trimeric chlorophyll a/c2-peridinin proteins in the dinoflagellate Symbiodinium

The chlorophyll a-chlorophyll c₂-peridinin-protein (apcPC), a major light harvesting component in peridinin-containing dinoflagellates, is an integral membrane protein complex. We isolated functional acpPC from the dinoflagellate Symbiodinium. Both SDS-PAGE and electrospray ionization mass spectrometry (ESI-MS) analysis quantified the denatured subunit polypeptide molecular weight (MW) as 18 kDa. Size-exclusion chromatography (SEC) and blue native gel electrophoresis (BN-PAGE) were employed to estimate the size of native acpPC complex to be 64–66 kDa. We also performed native ESI-MS, which can volatilize and ionize active biological samples in their native states. Our result demonstrated that the native acpPC complex carried 14 to 16 positive charges, and the MW of acpPC with all the associated pigments was found to be 66.5 kDa. Based on these data and the pigment stoichiometry, we propose that the functional light harvesting state of acpPC is a trimer. Our bioinformatic analysis indicated that Symbiodinium acpPC shares high similarity to diatom fucoxanthin Chl a/c binding protein (FCP), which tends to form a trimer. Additionally, acpPC protein sequence variation was confirmed by de novo protein sequencing. Its sequence heterogeneity is also discussed in the context of Symbiodinium eco-physiological adaptations.     

Mutations in cytochrome b that affect kinetics of the electron transfer reactions at center N in the yeast cytochrome bc1 complex

We have examined the pre-steady-state kinetics and thermodynamic properties of the b hemes in variants of the yeast cytochrome bc₁ complex that have mutations in the quinone reductase site (center N). Trp-30 is a highly conserved residue, forming a hydrogen bond with the propionate on the high potential b heme (b_H heme). The substitution by a cysteine (W30C) lowers the redox potential of the heme and an apparent consequence is a lower rate of electron transfer between quinol and heme at center N. Leu-198 is also in close proximity to the b_H heme and a L198F mutation alters the spectral properties of the heme but has only minor effects on its redox properties or the electron transfer kinetics at center N. Substitution of Met-221 by glutamine or glutamate results in the loss of a hydrophobic interaction that stabilizes the quinone ligands. Ser-20 and Gln-22 form a hydrogen-bonding network that includes His-202, one of the carbonyl groups of the ubiquinone ring, and an active-site water. A S20T mutation has long-range structural effects on center P and thermodynamic effects on both b hemes. The other mutations (M221E, M221Q, Q22E and Q22T) do not affect the ubiquinol oxidation kinetics at center P, but do modify the electron transfer reactions at center N to various extents. The pre-steady reduction kinetics suggest that these mutations alter the binding of quinone ligands at center N, possibly by widening the binding pocket and thus increasing the distance between the substrate and the b_H heme. These results show that one can distinguish between the contribution of structural and thermodynamic factors to center N function.