利用稀土位移探针对水溶液中L-精氨酸构象的~(13)C-NMR研究

采用重稀土离子(Dy、Ho、Er、Tm、Yb)研究了水溶液中L-精氨酸的构象。结果表明,距稀土配位中心4个或4个键以上的配体核的接触位移都很小,在稀土离子附近的配体核具有显著的接触位移。通过对配体磁性核结构因子的实验值进行模拟,建立了水溶液中L-精氨酸的整体构象。在L-精氨酸稀土配合物中,配体的羧基与稀土离子配位,配体的骨架结构位于稀土离子的零偶极位移锥面的外侧。对于羧基的双齿配位模式,计算得到的RE~(3+)-O键长为0.21nm。在溶液中配体以伸展状态存在,分子骨架呈全反式构象。     

硼酸对赖氨酸盐酸盐在半胱氨酸自组装膜上晶体生长的影响

研究了利用半胱氨酸自组装膜对诱导L-赖氨酸盐酸盐晶体生长的影响。实验发现,赖氨酸盐酸盐优先在水溶液内而不是在自组装膜表面成核生长为晶体,这是首次发现自组装单层对赖氨酸盐酸盐的成核有抑制作用。对自组装单层上的成核机理进行了探讨,2种氨基酸正电荷间的排斥作用可能是自组装膜抑制赖氨酸盐酸盐成核的原因。在赖氨酸盐酸盐溶液中加入了等摩尔硼酸,XRD结果表明,硼酸晶体优先生长在自组装单层上。而析出后的硼酸晶体又加速促进了赖氨酸盐酸盐的结晶析出。扫描电子显微镜表征发现,加入硼酸后赖氨酸盐酸盐晶体的形貌发生了很大变化,由原来的块状变为针状。实验结果表明,加入硼酸,可以加速赖氨酸盐酸盐的晶体生长并可以调控晶体的形貌和取向。     

L—抗坏血酸与稀土离子作用的NMR研究

利用稀土诱导位移方法研究了水溶液中L-抗坏血酸(Vc)与三价稀土离子的配位作用。在弱酸性条件下二者形成1:1的配合物,稳定常数为8.0M~(-1)。抗坏血酸通过内脂环3位碳上的羟基氧与稀土离子配位,RE~(3 )-O键长为2.0,内脂基和其他碳上的羟基不与稀土离子配位。在配合物中C(3)-C(4)-C(5)-C(6)成反式构象。C(5)-C(6)键与配合物的零偶极位移锥面交叉。     

用双谱仪方法和静态魔角方法研究稀土水溶液~1H 的化学位移

Reuben 等以叔丁醇作内标研究过稀土高氯酸盐水溶液~1H 的 NMR 信号,观察到较大的顺磁位移(如在1MDy~(3 )溶液中可达10ppm),其中以偶极位移贡献为主.为了避免稀土离子对内标的影响,在实验中常采用外标法.本文以 HDO 为外标,研究了稀土氯化物的水溶液,在0.1M 的稀土水溶液中观察到~1H 信号有相当大的位移(如在0.1MDy~(3 )溶液中高达20ppm,相当于在1M 浓度时为200ppm).用双谱仪方法和静态魔角方法进行研究后,确认此种位移现象系稀土水溶液各向同性的整体磁化率(bulk susceptibility)所致.由此可见,稀土水溶液~1H 化学位移所受到的整体磁化率的影响要远远大于顺磁位移效应.     

聚(L-丙氨酸)-聚羟乙基谷氨酰胺双嵌段两亲性共聚多肽的合成及其在水溶液中的自组装

利用光气法分别以L-谷氨酸和L-丙氨酸为原料,合成了γ-谷氨酸苄酯-NCA单体和L-丙氨酸-NCA单体,再以三乙胺为引发剂,合成了聚(L-丙氨酸)-聚(γ-谷氨酸苄酯)(PLA50-b-PBLG20)双嵌段共聚多肽,并用乙醇胺对其中的PBLG嵌段进行亲核取代,把疏水性的苄酯侧链变为亲水性的羟烷酰胺侧链,得到双亲性的聚(L-丙氨酸)-聚羟乙基谷氨酰胺(PLA-b-PHEGA)双嵌段共聚多肽.利用红外光谱和核磁共振谱对产物进行了表征,用TEM研究了双嵌段共聚多肽PLA50-b-PHEGA20在水溶液中的自组装.研究结果表明,双嵌段共聚多肽在水溶液中可自组装形成囊泡.     

L—赖氨酸锌配合物结构的理论研究

采用量子化学半经验PM3方法,优化了L-赖氨酸锌配合物的几何结构,对L-赖氨酸锌配合物的结构、电子性质进行了分析,结果表明：计算结果与实验得到的晶体结构基本吻合,证实了L-赖氨酸锌配合物中Zn具有五配位结构,同时对体系的净电荷分布及前线轨道成分进行了分析。     

荧光法研究氧氟沙星与牛乳铁蛋白的相互结合作用

用荧光光谱法研究了稀水溶液中牛乳铁蛋白与氧氟沙星分子之间的结合作用机制.以荧光猝灭法测定了该反应的结合常数K, 结合位点数n；并依据Förster偶极-偶极无辐射能量转移机制,得到氧氟沙星和牛乳铁蛋白色氨酸残基之间结合距离r和能量转移效率E；并用同步荧光技术考察了氧氟沙星对牛乳铁蛋白构象的影响；牛乳铁蛋白与氧氟沙星分子之间有较强的结合作用,而且氧氟沙星对牛乳铁蛋白的构象有一定的影响.     

固体核磁共振高速魔角旋转条件下对称性脉冲零量子同核重耦技术（英文）

基于偶极耦合的同核相关固体核磁共振实验广泛应用于结构表征,RFDR是其中最广泛使用的零量子同核重耦序列之一。此前的研究中证明了RFDR的重耦效率非常依赖于魔角旋转转速、共振偏置、射频场不均匀性、化学位移各项异性及其它多种因素。本文基于对称性序列原理,在中等和高转速下考察了一系列RN_N~1 (N≧4, N为偶数)对称性序列,并结合多种相位循环,以充分研究零量子重耦序列原理并实现均匀的宽带同核重耦。不同RN对称性序列的零量子偶极重耦效率进一步在~(13)C、~(15)N均匀标记的L-组氨酸以及微晶动力蛋白轻链(LC8)样品上得到检验。     

两亲刚性三嵌段聚多肽的合成及其自组装行为

通过胱胺三甲基硅氮烷(N-TMS)引发N-羧基内酸苷(NCA)开环聚合,合成了2种两亲刚性三嵌段聚多肽聚(L-赖氨酸-ε-端甲基二缩乙二醇酰胺)-b-聚(L-谷氨酸-γ-苄酯)-b-聚(L-赖氨酸-ε-端甲基二缩乙二醇酰胺)(P[Lys-(EG)_2]-b-PBLG-b-P[Lys-(EG)_2])和聚(L-谷氨酸-γ-苄酯)-b-聚(L-赖氨酸-端甲基二缩乙二醇酰胺)-b-聚(L-谷氨酸-γ-苄酯)(PBLG-b-P[Lys-(EG)_2]-bPBLG),并用其在N,N-二甲基甲酰胺(DMF)-水的混合溶液中制备了三嵌段聚多肽的自组装体。采用核磁共振氢谱(1 H-NMR)和凝胶渗透色谱(GPC)表征了三嵌段聚多肽的结构,通过透射电子显微镜(TEM)研究了三嵌段聚多肽在混合溶液中的自组装行为。结果表明:通过N-TMS引发NCA开环聚合得到的三嵌段聚多肽的分子量与其理论值基本一致,且分子量分布窄;聚多肽在DMF-水的混合溶液中分别形成了球状胶束、大复合胶束、棒状大复合胶束等组装体;其自组装行为与特殊的全刚性嵌段结构有关。     

三种氨基酸在尿素水溶液中的体积性质

精密密度法详细测定甘氨酸、L-丙氨酸、L-丝氨酸在尿素水溶液中的表观摩尔体积,计算了三种氨基酸从水到尿素水溶液的迁移偏摩尔体积,结合前期的氨基酸从水到尿素水溶液的迁移焓,探讨尿素水溶液的结构特点及其对氨基酸与尿素在水溶液中相互作用的影响。结果表明,尿素分子在水溶液中自缔合,引起溶剂结构的变化并削弱其与氨基酸分子的结构相互作用,造成氨基酸从水到尿素水溶液的迁移偏摩尔体积和迁移焓随尿素浓度的增加而出现多个变化点,这一效应随着氨基酸疏水性的增强而增大,表明氨基酸的疏水性越强,其与尿素相互作用引起的去水化作用越明显。     

Conformational study of L-lysine in aqueous solution by lanthanide shift probes

Conformation of L-lysine in aqueous solution was investigated by lanthanide shift probes (Dy, Ho, Er, Tm and Yb). Reilley's method was employed to separate the contact and dipolar components of the ¹³C paramagnetic shifts. This study reveals that Cα shift has the largest contact contribution while the other carbon shifts are dominated by the dipolar contribution. The average overall conformation of L-lysine in aqueous solution is extended with the molecular backbone in trans form. In the complex, lanthanide ion coordinates to the carboxyl group with Ln—O bond length 2.2 Å and the whole ligand is located outside the zero-dipolar shift cone of the lanthanide ion. The electronic spin density distribution on the ligand nuclei shows that the spin polarization is the predominant mechanism of the contact interaction for nuclei in close proximity to the bound lanthanide ion.     

Triple oscillating field technique for accurate distance measurements by solid-state NMR

We present a new concept for homonuclear dipolar recoupling in magic-angle-spinning (MAS) solid-state NMR experiments which avoids the problem of dipolar truncation. This is accomplished through the introduction of a new NMR pulse sequence design principle: the triple oscillating field technique. We demonstrate this technique as an efficient means to accomplish broadband dipolar recoupling of homonuclear spins, while decoupling heteronuclear dipolar couplings and anisotropic chemicals shifts and retaining influence from isotropic chemical shifts. In this manner, it is possible to synthesize Ising interaction (2IzSz) Hamiltonians in homonuclear spin networks and thereby avoid dipolar truncation--a serious problem essentially all previous homonuclear dipolar recoupling experiments suffer from. Combination of this recoupling concept with rotor assisted dipolar refocusing enables easy readout of internuclear distances through comparison with analytical Fresnel curves. This forms the basis for a new class of solid-state NMR experiments with potential for structure analysis of uniformly 13C labeled proteins through accurate measurement of 13C-13C internuclear distances. The concept is demonstrated experimentally by measurement of C alpha-C', C beta-C', and C gamma-C' internuclear distances in powder samples of the amino acids L-alanine and L-threonine.     

1H and 13C NMR investigation of the influence of nonligated residue contacts on the heme electronic structure in cyanometmyoglobin complexes reconstituted with centro- and pseudocentrosymmetric hemins

The 1H and 13C chemical shifts for the heme methyls of low-spin, ferric sperm whale cyanometmyoglobin reconstituted with a variety of centrosymmetric and pseudocentrosymmetric hemins have been recorded and analyzed to shed light on the nature of heme-protein contacts, other than that of the axial His, that modulate the rhombic perturbation to the heme's in-plane electronic asymmetry. The very similar 1H dipolar shifts for heme pocket residues in all complexes yield essentially the same magnetic axes as in wild type, and the resultant dipolar shifts allow the direct determination of the heme methyl proton and 13C contact shifts in all complexes. It is demonstrated that, even when the magnetic axes and anisotropies are known, the intrinsic uncertainties in the orientational parameters lead to a sufficiently large uncertainty in dipolar shift that the methyl proton contact shifts are inherently significantly less reliable indicators of the unpaired electron spin distribution than the methyl 13C contact shifts. The pattern of the noninversion symmetry in 13C contact shifts in the centro- or pseudocentrosymmetric hemes is shown to correlate with the positions of aromatic rings of Phe43(CD1) and His97(FG3) parallel to, and in contact with, the heme. These results indicate that such pi-pi interactions significantly perturb the in-plane asymmetry of the heme pi spin distribution and cannot be ignored in a quantitative interpretation of the heme methyl 13C contact shifts in terms of the axial His orientation in b-type hemoproteins.     

Evaluation of dipolar shifts and paramagnetic anisotropy in penta coordinated cobalt(II) complexes

The isotropic proton shifts for the pyridine N-oxide and γ-picoline N-oxide protons have been observed in the penta coordinated adducts of these bases with bis[di(p-tolyl)dithiophosphinato] cobalt(II). The contribution to the observed shifts due to dipolar interaction has been calculated. From the dipolar shifts, it was ascertained that the pyridine N-oxide complexes have a bent structure in solution with a Co-O-N angle of 125°. An estimate of the paramagnetic anisotropy of the cobalt complex yields K_?-K_⊥ = 4244 × 10^?6 cm³/mole.     

水溶液中Ln(Ⅱ)-TTHA络合物的多核NMR研究

通过对11种镧系-三乙烯四胺六乙酸络合物诱导的²³Na、²H、¹⁷O和³⁵Cl的化学位移测定,研究了该系列络合物的位移试剂性质。结果表明,轴对称的Ln(TTHA)^3-络合物是较好的水溶性位移试剂,尤其是²³Na位移与理论偶极位移一致,表明此络合物是最好的正电荷底物之一。     

The dependence of reversible potentials on the form of modification of edge plane pyrolytic graphite electrodes in voltammetric studies on rubredoxin and ferredoxin from Clostridium pasteurianum

Reversible voltammetry of the anionic rubredoxin and ferredoxin proteins at a pyrolytic graphite edge plane electrode can be observed either by ex situ modification of the electrode with multivalent cations (poly(L-lysine), Cr(III) complexes) or by addition of these cations to the solution. However, poly(L-lysine) binds irreversibly to pyrolytic graphite and voltammograms obtained at an ex situ form of this modified electrode provide convenient reference points for estimation of the thermodynamic significance of solution phase interactions between the cationic modifiers and the proteins. Results obtained show that significant shifts in reversible potential are induced by the presence of free modifier in solution. Examination of the recombinant proteins and certain mutant forms suggests that association of the anionic proteins with cationic modifiers in solution is the primary cause of this shift. The effect of background ionic strength is minor in comparison. The conclusion is that the modifiers added to the solution phase are not thermodynamically innocent.     

Heteronuclear dipolar decoupling effects on multiple-quantum and satellite-transition magic-angle spinning NMR spectra

We here report on the influence of heteronuclear dipolar decoupling on the (27)Al 3QMAS, 5QMAS, and the double-quantum filter-satellite-transition magic-angle spinning (DQF-STMAS) spectra of a strongly dipolar-coupled system, gibbsite. The requirements for heteronuclear dipolar decoupling increase with the order of coherence evolving in the indirect dimension of a two-dimensional (2D) experiment. The isotropic line width of the high-resolution 2D spectra, in samples like gibbsite, is composed of four parts: the distribution of isotropic shifts (delta(ISO), delta(QIS)), the homogeneous broadening related to the proton-proton flip-flop terms, the (27)Al-(27)Al homonulcear dipolar couplings, and the (1)H-(27)Al heteronuclear dipolar couplings. It is shown that, even in the case of gibbsite, where a strong proton-proton bath exists, the main resolution limiting factor in these experiments resides in the (1)H-(27)Al dipolar interaction.     

1H and 13C NMR studies of lanthanide complex with 5-sulfosalicylic acid

In this paper, paramagnetic shifts have been measured for all ¹H and ¹³C nuclei of 5-sulfosalicylic acid (SSA) in the presence of the lanthanide ions in the second half of the series. The ligand forms isostructural complexes with these ions in aqueous solution. The separation of LISs was carried out by the use of the Reilley method and the calculated dipolar shifts were used to simulate the coordination structure of the complex. The result reveals that SSA is coordinated to lanthanide ion via two oxygens, one from the carboxylic group and the other from the phenolic group with Ln–O bond lengths equal to 2.47 Å. The lanthanide ion lies on the benzene plane and the carboxylic group is twisted 20° from the benzene ring. Of all the nuclei examined, those in the six-membered chelate ring experience significant dipolar interactions and contact interactions. Small |G/A| ratios were obtained for two protons five bonds away from the central lanthanide ion, which shows that the number of bonds alone cannot be used as a criterion for neglecting contact shifts in aromatic ligand.     

NMR “Crystallography” for Uniformly (13C, 15N)-Labeled Oriented Membrane Proteins

In oriented-sample (OS) solid-state NMR of membrane proteins, the angular-dependent dipolar couplings and chemical shifts provide a direct input for structure calculations. However, so far only ¹H–¹⁵N dipolar couplings and ¹⁵N chemical shifts have been routinely assessed in oriented ¹⁵N-labeled samples. The main obstacle for extending this technique to membrane proteins of arbitrary topology has remained in the lack of additional experimental restraints. We have developed a new experimental triple-resonance NMR technique, which was applied to uniformly doubly (¹⁵N, ¹³C)-labeled Pf1 coat protein in magnetically aligned DMPC/DHPC bicelles. The previously inaccessible ¹H_α–¹³C_α dipolar couplings have been measured, which make it possible to determine the torsion angles between the peptide planes without assuming α-helical structure a priori. The fitting of three angular restraints per peptide plane and filtering by Rosetta scoring functions has yielded a consensus α-helical transmembrane structure for Pf1 protein.