Structure and function of haemoglobin philly (Tyr C1 (35) β→Phe)

We have purified haemoglobin Philly by isoelectric focusing on polyacrylamide gel, and studied its oxygen equilibrium, proton nuclear magnetic resonance spectra, mechanical stability, and pH-dependent u.v. difference spectrum. Stripped haemoglobin Philly binds oxygen non-co-operatively with high affinity. Inorganic phosphate and 2,3-diphosphoglycerate have little effect on the equilibrium curve, but inositol hexaphosphate lowers the affinity and induces co-operativity. These properties are explained by the nuclear magnetic resonance spectra which show that stripped deoxyhaemoglobin Philly has the quaternary oxy structure and that inositol hexaphosphate converts it to the deoxy structure. An exchangeable proton resonance at ?8.3 p.p.m. from water, which is present in oxy- and deoxyhaemoglobin A, is absent in both these derivatives of haemoglobin Philly and can therefore be assigned to one of the hydrogen bonds made by tyrosine C1-(35)β, probably the one to aspartate H8(126)α at the α₁β₁ contact. Haemoglobin Philly shows the same pH-dependent u.v. difference spectrum as haemoglobin A, only weaker, so that a tyrosine other than 35β must be mainly responsible for this.     

A new missense mutation (Cys297→Phe) of the low density lipoprotein receptor in Italian patients with familial hypercholesterolemia (FHTrieste)

During a survey of the mutations of the low density lipoprotein receptor (LDL-R) gene in Italian patients with familial hypercholesterolemia (FH), we identified a novel point mutation, that creates a new EcoRI site at the 5 end of exon 7, in a heterozygous FH subject (FH-100). The sequence of a cDNA fragment encompassing exon 7 showed the presence of a GT transversion in codon 297; this created a new EcoRI site and produced a missense mutation, leading to a Cys₂₉₇Phe substitution in repeat A of the epidermal growth factor (EGF) precursor homology domain of LDL-R. Since the substitution of Cys₂₉₇ disrupts the intracellular transport of the LDL-R protein, as previously demonstrated by site-directed mutagenesis, we suggest that this mutation is the cause of FH in the FH-100 proband. We screened the DNA of 303 Italian FH patients by amplification of exon 7 from genomic DNA followed by digestion with EcoRI or by Southern blotting. Two individuals (FH-64 and FH-127) were found to be carriers of the Cys₂₉₇Phe mutation. Restriction fragment length polymorphism analysis demonstrated that, in two kindreds (FH-64 and FH-100), the haplotype in linkage with the Cys₂₉₇Phe mutation was the same, suggesting the presence of a common ancestor. The Cys₂₉₇Phe mutation has been designated FH_Trieste after the name of the city in Northern Italy from which probands FH-100 and FH-127 originate.     

TheX → Y → Z scheme after 23 years

Mathematical modelling of the course of the immune response is undoubtedly one of the most progressive and most promising areas of modern immunology. Mathematical models (along with computer programs) can be taken as "the only means of thoroughly testing and examining a large and intricate theory" (Partridge et al. 1984). The first phase of construction of mathematical models is the formulation of assumptions based on the knowledge of the facts to be modelled (manifested usually in a scheme of the presumed course of the modelled process). The first mathematical models of immune response were based on the hypothesis of a two-stage differentiation of cells participating in the humoral response, published in Prague 23 years ago (Sercarz and Coons 1962; Sterzl 1962) and illustrated by the X----Y----Z scheme. Many contemporary mathematical models still stem from this scheme which undoubtedly fits the fundamental data concerning the immune system.     

Total synthesis,purification, and characterization of human [Phe(p-CH2SO3Na)52, Nle32,53,56, Nal55]-CCK20–58, [Tyr52, Nle32,53,56,Nal55]-CCK-58, and [Phe(p-CH2SO3Na)52, Nle32,53,56, Nal55]-CCK-58

The synthesis of [Phe(p-CH₂SO₃Na)⁵², Nle^32,53,56 Nal⁵⁵]-CCK_20–58, [Tyr⁵², Nle^32,53,56, Nal⁵⁵]-CCK-58 and of [Phe(p-CH₂SO₃Na)⁵², Nle^32,53,56, Nal⁵⁵]-CCK-58 using the (9-fluorenylmethyloxy)-carbonyl (Fmoc) strategy on a 2,4-DMBHA resin is described. The crude peptide preparations were extremely complex when analyzed by RP-HPLC, capillary zone electrophoresis (CZE), and ion-exchange chromatography (IE-FPLC). We found that the most effective strategy for purification included cation-exchange chromatography followed by a RP-HPLC desalting step. The highly purified peptides (purity greater than 90%) were characterized by RP-HPLC, size exclusion HPLC (SEC), IE-FPLC, CZE, mass spectrometry, amino acid analysis, and Edman sequence analysis {for [Tyr⁵², Nle^32,53,56, Nal⁵⁵]-CCK-58}. The results demonstrate the applicability of the 2,4-DMBHA resin for Fmoc solid-phase synthesis of long peptides amides (58 residues in length in this case) as well as the efficacy of an FPLC/RP-HPLC approach for the purification of very long, heterogeneous crude peptides, allowing a true assessment of the biological properties of these analogs to be carried out. [Phe(p-CH₂SO₃Na)⁵², Nle^32,53,56, Nal⁵⁵]-CCK_20–58 was less than 1% as potent as CCK-8 while [Tyr⁵², Nle^32,53,56, Nal⁵⁵]-CCK-58 and [Phe(p-CH₂SO₃Na)⁵², Nle^32,53,56, Nal⁵⁵]-CCK-58 were inactive at the doses tested (<0.01%).     

“脊髓→皮层→脊髓”环路易化触觉信息传递

正在躯体感觉研究领域,人们长期关注从外周到脊髓再到大脑,这样一条由下至上(bottom up)的传递通路,而对于来自大脑尤其是皮层发出的由上至下(top down)的感觉调控通路研究还比较少。2018年9月哈佛医学院何志刚和Clifford J.Woolf教授以及美国国立卫生研究院Wang Hongkuan教授团队共同在Nature杂志发表论文,指出躯体感觉皮层可通过皮质脊髓束     

在教学过程中贯穿激发→实现→强化学习动机的探索

学生动机是指推动学生学习的内在力量,是学生学习的推动者。课堂教学中学习动机分为激发、实现、强化3个阶段。因此,在教学过程中应贯穿激发学习动机一实现学习动机一强化学习动机,从而提高教学效果。l动机的激发阶段1．l有趣的导言或提出令人迷惑不解的矛盾问题激发学生的学习动机。例如：教师说："鸟为什么能飞？也许你会回答,因为鸟有翅膀,那么在人的双臂上缚上人工的翅膀人能不能像鸟一样S翔呢？"学生产生需要知道的心理,而要彻底明白这个道理,他们必须认真去听课。又例如在讲无机盐对植物的作用一节,教师可先讲一个小故事："…     

Glu–Phe from onion (Allium Cepa L.) attenuates lipogenesis in hepatocytes

A Glu–Phe (EF) was isolated from onion (Allium cepa L. cv. Sunpower). The chemical structure of EF was determined by nuclear magnetic resonance and electrospray ionization–mass (ESI?MS) spectroscopy. We showed that EF reduced lipid accumulation in mouse hepatocytes by inhibiting the expression of sterol regulatory element-binding protein-1c (SREBP–1c) and its lipogenic target genes. We also found that AMP-activated protein kinase (AMPK) was required for the inhibitory effect of EF on lipid accumulation in mouse hepatocytes. Furthermore, EF was qualified in nine onion cultivars by selective multiple reaction-monitoring detection of liquid chromatography–ESI?MS. These results suggest that EF could contribute to the beneficial effect of onion supplement in maintaining hepatic lipid homeostasis.     

Induction of T → G and T → A transversions by 5-formyluracil in mammalian cells

Oxidatively damaged thymine, 5-formyluracil (5-fU), was incorporated into a predetermined site of double-stranded shuttle vectors. The nucleotide sequences in which the modified base was incorporated were 5′-CFTAAG-3′ and 5′-CTFAAG-3′ (F represents 5-fU), the recognition site for the restriction enzyme AflII (5′-CTTAAG-3′). The 5-fU was incorporated into a template strand of either the leading or lagging strand of DNA replication. The modified DNAs were transfected into simian COS-7 cells, and the DNAs replicated in the cells were recovered and were analyzed after the second transfection into Escherichia coli. The 5-fU did not block DNA replication in mammalian cells. The 5-fU residues were weakly mutagenic, and their mutation frequencies in double-stranded vectors were 0.01–0.04%. The T → G and T → A transversions were the mutations found most frequently, suggesting the formation of 5-fU·C and 5-fU·T base pairs, respectively. This is the first report that clearly shows the induction of transversion mutations by an oxidized pyrimidine base in DNA in mammalian cells.     

B → A → B Transitions in a Molecular Dynamics Trajectory of Low Salt DNA Solution

A molecular dynamics simulation of the DNA dodecamer d(CGCATATATGCG) has been performed with AMBER 5.0 under low salt conditions. Both B A and A B transitions are observed. This may have biological significance for the formation of complexes between DNA and TATA-box binding proteins.Supplementary material to this paper is available in electronic form at http://dx.doi.org/10.1007/s0089400060654     

"应用→原理→应用创新"教学模式在高中生物学复习教学中的尝试

“应用→原理→应用创新”教学模式,强调在培养广大普通劳动者的基础上继续培养创新型人才,这不但有利于提高学生的学业成绩,同时也有利于提高学生的学习兴趣、培养动手和实践能力等。以高中生物学复习教学为例,简单介绍了本教学模式的具体应用及其效果。     

小角中子散射研究铯对溶液中tRNA~(Phe)构象的影响

用小角中子散射技术研究生物大分子的构象,图样清晰,可同时获得结构参数和热力学参数。 tRNA~(phe)在pH7.0的CsCl-TMA溶液中会形成Cs-tRNA~(phe),由于溶液中的电荷颗粒的作用使tRNA~(phe)的构象可能发生变化,本文研究不同浓度CsCl-TMA溶液中Cs对tRNA~(phe)在溶液中构象有无变化。 制备各浓度的CsCl-TMA—D_2O-tRNA~(phe)和以H_2O代D_2O的上述溶液样品。在小角中子散射装置D_(11)上测定Cs-tRNA~(qhe)的结构参数和热力学参数。 实验结果是不同浓度的CsCl-TMA-D_2O溶液中的Cs-tRNA~(phe)迥旋半径Rg的数值无大变化,中子散射密度增量(ds/dc)~2即I(o)/C的数值大致相同,颗粒的微分体积V_2值随盐浓度增加稍有加大,体积参数φ(?)的绝对值随盐浓度稍加.从以上结构参数Rg和热力学参数dρ/dc,V_2和φ′s等值分析发现Cs-tRNA~(phe)在Cs的作用下在溶液中构象无大变化。##原图像不清晰     

Enhanced β-turn conformational stability of tripeptides containing ΔPhe in cis over trans configuration

Conformations of three pairs of dehydropeptides with the opposite configuration of the ΔPhe residue, Boc-Gly-Δ^Z/EPhe-Phe-p-NA (Z- p -NA and E- p -NA), Boc-Gly-Δ^Z/EPhe-Phe-OMe (Z-OMe and E-OMe), and Boc-Gly-Δ^Z/EPhe-Phe-OH (Z-OH and E-OH) were compared on the basis of CD and NMR studies in MeOH, TFE, and DMSO. The CD results were used as the additional input data for the NMR-based calculations of the detailed solution conformations of the peptides. It was found that Z- p -NA, E- p -NA, Z-OMe, and Z-OH adopt the β-turn conformations and E-OMe and E-OH are unordered. There are two overlapping type III β-turns in Z- p -NA, type II’ β-turn in E- p -NA, and type II β-turn in Z-OMe and Z-OH. The results obtained indicate that in the case of methyl esters and peptides with a free carboxyl group, Δ^ZPhe is a much stronger inducer of ordered conformations than Δ^EPhe. It was also found that temperature coefficients of the amide protons are not reliable indicators of intramolecular hydrogen bonds donors in small peptides.     

金属内肽酶底物Glutaryl－Ala－Ala－Phe－2NA的合成

文本以苯丙萘胺（Ｐｈｅ－２ＮＡ）为原料,通过Ｂｏｃ－Ｎ－羟基丁二酰亚胺活化酯法合成了金属内肽酶底物Ｇｌｕｔａｒｙｌ－Ａｌａ－Ａｌａ－Ｐｈｅ－２ＮＡ     

Characterization of the RcsC→YojN→RcsB Phosphorelay Signaling Pathway Involved in Capsular Synthesis in Escherichia coli

Escherichia coli and other enteric microorganisms produce an extracellular polysaccharide capsule, called colanic acid, under certain environmental conditions. This capsular synthesis is regulated by the RcsC (sensor kinase)→YojN (phosphotransfer intermediate)→RcsB (response regulator) phosphorelay signal transduction under certain growth conditions. Nonetheless, little is known about signals that exaggerate the Rcs-system. To gain insight into signals that activate the Rcs-system, here we searched for genes that activate the Rcs-system, provided that those on a multicopy plasmid were introduced into E. coli. We identified several such genes, namely, rcsB, rcsA, djlA, lolA, and ompG. The DjlA, LolA, and OmpG proteins are particularly interesting in that they are all located on the cell surface, where the primary sensor RcsC histidine-kinase is localized. Implications of these findings are discussed with special reference to the mechanism by which RcsC perceives external signals.     

Functional analysis of the GAF domain of NifA in <Emphasis Type="Italic">Azospirillum brasilense</Emphasis>: effects of Tyr→Phe mutations on NifA and its interaction with GlnB

Regulation of NifA activity in Azospirillum brasilense depends on GlnB (a PII protein), and it was previously reported that the target of GlnB activity is the N-terminal domain of NifA. Furthermore, mutation of the Tyr residue at position 18 in the N-terminal domain resulted in a NifA protein that did not require GlnB for activity under nitrogen fixation conditions. We report here that a NifA double mutant in which the Tyr residues at positions 18 and 53 of NifA N-were simultaneously replaced by Phe (NifA-Y1853F) displays high nitrogenase activity, which is still regulatable by ammonia, but not by GlnB. The yeast two-hybrid technique was used to investigate whether GlnB can physically interact with wild-type and mutant NifA proteins. GlnB was found to interact directly with the N-terminal GAF domain of wild-type NifA, but not with its central or C-terminal domain. GlnB could still bind to the single NifA mutants Y18F and Y53F. In contrast, no interaction was detected between GlnB and the double mutant NifA-Y18/53F or between GlnB and NifA-Y43.     

Reverse Relationship between α-Carbon Chirality and Helix Handedness in (αMe)Phe Peptides

Abstract

The crystal-state preferred conformations of two tripeptides, one tetrapeptide, and one pen- tapeptide, each containing a single residue of the chiral, C^α,α-disubstituted glycine C^α-methyl, C^α-benzylglycine [(αMe)Phe], have been determined by X-ray diffraction. The tripeptides are Z-L-(αMe)Phe-(Aib)₂-OH dihydrate and Z-Aib-D-(αMe)Phe-Aib-OtBu, the tetrapeptide is Z-(Aib)₂-D-(αMe)Phe-Aib-OtBu, and the pentapeptide is pBrBz-(Aib)₂-DL-(αMe)Phe-(Aib)₂-OtBu. While the two tripeptides are folded in a β-bend conformation, two such conformations are consecutively formed by the tetrapeptide. The pentapeptide adopts a regular 3₁₀-helix promoted by three consecutive β-bends. This study confirms the strong propensity of short peptides containing C^α-methylated α-aminoacids to fold into β-bends and 3₁₀-helical structures. Since Aib is achiral, the handedness of the observed bends and helices is dictated by the presence of the (αMe)Phe residue. In general, we have found that the relationship between (αMe)Phe chirality and helix handedness is opposite to that exhibited by protein aminoacids. A comparison with the preferred conformation of other extensively investigated C^α-methylated aminoacids is made.     

De novo trisomy 4pter→q21

Summary Clinical and cytogenetic findings in an infant girl with multiple congenital anomalies, principally anophthalmia, are presented. The patient's karyotype was 47,XX, +del(4)(pterq21), the largest partial trisomy of chromosome 4 reported. The possible mechanism of the origin of this abnormality is discussed.To whom offprint requests should be sent     

“de novo” trisomy 1q32→1qter and monosomy 3p25→3pter

Summary The syndrome of familial lymphedema (type Meige) with distichiasis was observed in father and son. The association with uvula bifida and submucous cleft of the palate is described for the first time.     

Probe design and synthesis of Galβ(1→3)[NeuAcα(2→6)]GlcNAcβ(1→2)Man motif of N-glycan

Synthesis and clusterization of Galβ(1→3)[NeuAcα(2→6)]GlcNAcβ(1→2)Man motif of the N-glycan, as the molecular probes for their biological evaluation, are reported. Key step is the quantitative and the completely α-selective sialylation of the C5-azide N-phenyltrifluoroacetimidate with the disaccharide acceptor, Galβ(1→3)GlcNTroc. Clusterization of the 16 molecules of trisaccharide motif was also achieved by the ‘self-activating click reaction’. These probes could efficiently be labeled by biotin and/or other fluorescence- or radioactive reporter groups through either cross metathesis, acylation, Cu(I)-mediated Huisgen [2+3]-cycloaddition, or the azaelectrocyclization to utilize the various biological techniques.