慢性肺心病血浆游离氨基酸分析

测定了64例慢性肺心病急性加重期患者的17种血浆游离氨基酸含量。结果显示:肺心病和肺性脑病氨基酸总量明显低于正常对照组(P均<0.01);肺性脑病血浆支链氨基酸与芳香族氨基酸比值(BCAA/AAA)明显下降,与正常人和肺心病(不含肺性脑病)组相比有显著差异(P均<0.01)。提示,肺心病急性加重期存在有明显的氨基酸代谢障碍;肺性脑病的BCAA/AAA比值下降与肝性脑病时表现相似,推测氨基酸代谢率乱可能参与肺脑的发病过程。     

肝硬化时蛋白质与氨基酸代谢的改变及其临床意义

肝硬化时,机体多处于分解代谢状态,蛋白质合成虽有增加,但仍相对不足。肝硬化患者多有较好的节氮效应。肝硬化时血浆芳香族氨基酸谱(AAA)上升,支链氨基酸(BCAA)和BCAA/AAA 比值减低。肝硬化时可能有BCAA 转运系统的调节异常及分布异常。AAA 水平取决于肝脏损害的程度。AAA 水平高常伴有负氮平衡,BCAA/AAA 比值是判断肝脏损害程度的一个敏感指标。血浆氨基酸谱改变与肝性脑病的程度不相关。     

肝病患者氨基酸代谢紊乱的意义及其治疗

许多临床观察和动物实验提示,肝性脑病和多种肝病时氨基酸代谢有明显改变,主要表现为血液中芳香族氨基酸(AAA)如苯丙氨酸和酪氨酸的增多以及支链氨基酸(BCAA)如缬氨酸、亮氨酸和异亮氨酸的减少。BCAA 减少及/或 AAA 增多导致 BCAA/AAA 克分子比值降低。与此同时,蛋氨酸和半胱氨酸等     

静脉滴注支链氨基酸治疗肝性脑病是否有效?一项多中心研究

本文报道法国、瑞典五个医疗中心用支链氨基酸(BCAA)治疗肝性脑病研究的结果。采用BCAA溶液(20g/L,亮氨酸占70%,缬氨酸20%,异亮氨酸10%,加于5%糖水内)治疗,并以5%糖水为对照。方法:选酒精性、原发性胆汁性、隐原性与肝炎后性肝硬化和急性肝性脑病的患者共50例,内中2例Ⅱ级,18例Ⅲ级,30例Ⅳ_a～Ⅳ_c级,按随机双盲法分为两组,BCAA和安慰剂组各25例,后一组年龄稍轻。两组的肝性脑病程度分级、胃肠道出血的人数、输血量     

表皮生长因子(EGF)水平在慢性肝病中的动态变化及意义

目的观察慢性肝炎肝硬化患者血清表皮生长因子(EGF)及甘草酸二胺注射液治疗前后血清EGF含量变化.方法52例肝病患者分为3组.其中急性肝炎15例,慢性肝炎23例,肝炎后肝硬化14例,正常对照30例.EGF检测采用放射免疫分析方法.结果急性肝炎组、慢性肝炎组、肝炎后肝硬化组患者血清EGF水平与对照组相比具有显著性差异(P＜0.01),且在药物治疗前后其水平具有显著性差异,治疗后EGF水平明显低于治疗前(P＜0.01).相关性分析表明,肝功能中丙氨酸氨基转移酶与EGF呈正相关.讨论EGF可作为衡量肝脏炎症状况的参考指标.动态观察EGF水平对了解肝脏炎症及肝纤维化状况、疗效判断具有重要的参考价值.     

重型肝炎患者外周血干细胞因子水平及临床意义

目的: 探讨重型肝炎患者外周血干细胞因子(stem cell factor,SCF)与其肝损害程度及预后的关系.方法: 贵州省遵义医学院第一附属医院住院的肝炎患者45例及门诊体检者15例.肝炎患者分为急性肝炎组15例,慢性肝炎组16例和重型肝炎组14例;重型肝炎患者又分为好转组4例,死亡组10例.ELISA法检测各组血清SCF水平.结果: 重型肝炎患者血清SCF水平明显高于急性肝炎、慢性肝炎和健康对照组,差异具有统计学意义(2403.1±42.8 ng/L vs 2354.9±19.0ng/L,2376.7±16.4 ng/L,2358.4±16.0 ng/L,均P＜0.05).重型肝炎患者死亡组血清SCF水平明显高于好转组,差异有统计学意义(2418.1±50.7 ng/L vs 2376.3±11.7 ng/L,P＜0.05).血清SCF与肝细胞生长因子有明显正相关(r =0.38,P＜0.01).SCF与血清白蛋白、凝血酶原时间活动度呈明显负相关( P＜0.01).结论: 重型肝炎患者外周血清SCF水平随着肝损害程度加重而明显升高,提示严重肝损害时,肝再生可能需要干细胞的参与.     

MR化学位移成像脑代谢物改变与肝硬化的临床相关性

目的利用MRS评价肝硬化患者脑代谢物的异常变化及其与Child-Pugh分级、肝功指标的相关性。方法应用3.0T磁共振对52例乙型肝炎肝硬化患者(Child-Pugh A级患者20例、Child-Pugh B级患者14例、Child-Pugh C级患者18例)及15名慢性乙型肝炎患者(对照组)进行磁共振波谱分析,计算脑代谢物峰下面积Cho(胆碱)、Mi(肌醇)、NAA(N-乙酰天门冬氨酸)、Cr(肌酐)、Glx6(谷氨酰胺复合物)、脑代谢物峰下面积与Cr比值(NAA/Cr、Cho/Cr、Mi/Cr、Glx6/Cr)。结果①Child-Pugh A级组与对照组Mi、Glx6比较,差异有统计学意义。Child-Pugh B级组与Child-Pugh A级组Cho、Mi、Glx6比较,差异有统计学意义(P<0.01);Child-Pugh C级组与Child-Pugh B级组Cho、Mi比较,差异有统计学意义(P<0.01或P<0.05)。②Child-Pugh A级组与对照组Mi/Cr、Glx6/Cr比较,差异有统计学意义(P<0.01);Child-Pugh B组级与Child-Pugh A级组Cho/Cr、Mi/Cr、Glx6/Cr比较,差异有统计学意义(P<0.01或P<0.05);Child-Pugh C级组与Child-Pugh B级组Cho/Cr、Mi/Cr比较,差异有统计学意义(P<0.05)。③肝性脑病(HE)组与非肝性脑病组脑代谢物峰下面积比值Cho/Cr、Mi/Cr、Glx6/Cr比较,差异有统计学意义(P<0.01)。④Child-Pugh分级与Glx6/Cr呈正相关,与Cho/Cr、Mi/Cr呈负相关。⑤肝硬化患者TBil、PT与Cho/Cr、Mi/Cr呈负相关,与Glx6/Cr呈正相关。ALB与Cho/Cr、Mi/Cr呈正相关,与Glx6/Cr呈负相关。ALT与NAA/Cr、Cho/Cr、Mi/Cr、Glx6/Cr无明显相关性。结论肝硬化及肝性脑病患者存在脑代谢物浓度异常改变,可作为早期诊断肝硬化、肝性脑病及评价肝硬化、肝性脑病严重程度的一项指标。     

肝硬化时血浆支链氨基酸水平降低的机制

肝硬化时血浆氨基酸改变的特点是芳香族氨基酸(AAA)升高,支链氨基酸(BCAA)降低;这种改变的重要性在于它和肝性脑病有密切的关系.用富含BCAA的氨基酸液治疗肝性脑病的效益已为许多作者证实.因此,阐明血浆BCAA降低的机制和设计更合理的治疗有重要的意义.芳香族氨基酸主要在肝脏内代谢.肝硬化时血浆AAA水平的升高是由于肝清除能力减低之故.但支链氨基酸主要在外周组织内代谢,其降低与肝脏的代谢无关,而由下列多种因素引起.营养不良和饥饿肝病患者常因食欲不振、饮食习惯不良或有其它合并疾病而经常处于"饥饿"状态.长期饥饿使BCAA氧化供能增加,血浆BCAA降     

血清游离氨基酸检测在肝病中的临床应用

自1965年国外学者发现急、慢性肝病时血清氨基酸浓度有明显变化以后。Fischer于1974年在动物实验的基础上,应用特殊氨基酸溶液治疗11例肝性脑病患者取得了较好效果。并提出了以支/芳比值(BCAA/AAA)和肝功损害程度及其预后有密切关系的报告。近年来,国内外对肝昏迷、重症肝炎、肝硬化等肝病所引起的血清氨基酸代谢失衡学说以及根据这一理论开创的氨基酸治疗也日益受到重视。目前随着氨基酸谱     

重型肝炎血氨检测的价值的研究

目的 了解重型肝炎血氨特点及其临床价值.方法 用DRI-CHEM100型血氨分析仪检测40例重型肝炎患者、20例慢性肝炎患者及20名正常人的血清样本中的血氨水平.结果 重型肝炎患者血氨水平显著高于慢性肝炎患者(P<0.01);慢性肝炎患者血氨水平显著高于健康人(P<0.01).合并肝性脑病的重型肝炎患者血氨水平较无肝性脑病的重型肝炎患者的血氨水平略高,但无显著性差异(P>0.05).治愈、好转的重型肝炎患者与死亡、离院的重型肝炎患者治疗前血氨水平无显著性差异(P>0.05).结论 血氨检测对于诊断重型肝炎有一定价值.     

口服支链氨基酸治疗肝硬化低蛋白血症多中心总结

为验证口服支链氨基酸治疗肝硬化低蛋白血症的疗效,将149例患者随机分为两组,A组口服支链氨基酸,B组口服复方氨基酸胶囊,疗程9周。结果表明:A组血清白蛋白水平由治疗前(32.25±3.11)g/dl升高到(37.61±5.13)g/dl(P<0.05);B组血清白蛋白由治疗前(32.12±2.04)g/dl升高到(36.64±4.66)g/dl(P<0.05),两组对比,支链氨基酸组血清白蛋白升幅明显高于对照组。两组均未发生严重不良反应。因此,支链氨基酸是一种安全有效的提升肝病患者白蛋白水平的药品。     

Structure of a tRNA-dependent kinase essential for selenocysteine decoding

Compared to bacteria, archaea and eukaryotes employ an additional enzyme for the biosynthesis of selenocysteine (Sec), the 21^st natural amino acid (aa). An essential RNA-dependent kinase, O-phosphoseryl-tRNA^Sec kinase (PSTK), converts seryl-tRNA^Sec to O-phosphoseryl-tRNA^Sec, the immediate precursor of selenocysteinyl-tRNA^Sec. The sequence of Methanocaldococcus jannaschii PSTK (MjPSTK) suggests an N-terminal kinase domain (177 aa) followed by a presumed tRNA binding region (75 aa). The structures of MjPSTK complexed with ADP and AMPPNP revealed that this enzyme belongs to the P-loop kinase class, and that the kinase domain is closely related to gluconate kinase and adenylate kinase. ATP is bound by the P-loop domain (residues 11–18). Formed by antiparallel dimerization of two PSTK monomers, the enzyme structure shows a deep groove with positive electrostatic potential. Located in this groove is the enzyme''s active site, which biochemical and genetic data suggest is composed of Asp-41, Arg-44, Glu-55, Tyr-82, Tyr-83, Met-86, and Met-132. Based on structural comparison with Escherichia coli adenylate kinase a docking model was generated that assigns these amino acids to the recognition of the terminal A76-Ser moieties of Ser-tRNA^Sec. The geometry and electrostatic environment of the groove in MjPSTK are perfectly complementary to the unusually long acceptor helix of tRNA^Sec.     

生物工程合成生长激素对生长激素缺乏性侏儒血浆氨基酸谱的影响

本文对10例生长激素缺乏性侏儒症(GHD)患儿在应用生物工程合成生长激素(R-hGH)治疗初始及治疗3个月时的血浆游离氨基酸(FAA)谱进行了配对分析。治疗初始及治疗3个月时,注射R-hGH后4小时的血浆FAA中,半数以上的必需氨基酸(EAA)和大部分(8/12)非必需氨基酸(NEAA)显著减少,提示氨基酸参与了蛋白质合成及生糖生酮。治疗3个月时血浆大部分氨基酸于注射后4小时显著减少的程度不及治疗初始,可能为机体对较长时间hGH治疗的代谢适应。与治疗初始比较,R-hGH治疗3个月时间的空腹血浆FAA总值和尿素显著低下,提示机体氮排出减少和保留增加,多用于合成NEAA。甘氨酸、脯氨酸、谷氨酸等明显低下,说明骨组织在较高时间R-hGH作用下对氨基酸需求增加及组织之生长旺盛,亦提出应加强蛋白营养以满足生长需要。     

Alterations in carrier-mediated glutamine transport after a model of canine jejunal autotransplantation

The effects of small bowel transplantation (SBTx) on absorptive function are unknown. Preliminary experiments showed a decrease in absorption of glutamine. Our aim was to determine mechanisms of decreased ileal transport of glutamine utilizing a model of intestinal autotransplantation. Seven dogs were studied before and after a model of jejunoileal autotransplantation.In vivo absorption experiments were performed before and two and eight weeks postoperatively with an electrolyte solution containing glutamine (20 mM).In vitro glutamine transport was studied using brush-border membrane vesicles (BBMV) prepared from ileal mucosa obtained from six other dogs and compared to a controls.In vivo net absorptive flux of glutamine decreased at two weeks but returned toward baseline by eight weeks (P=0.06). Transport of glutamine into BBMVs was decreased at two weeks and remained decreased at eight weeks. , a measure of carrier affinity was unchanged but , a function of the number of transporter was decreased at two and eight weeks. Glucose transport was unchanged. It is concluded that jejunoileal autotransplantation decreases ileal absorption of glutamine by a decrease in carrier-mediated transport of glutamine.Supported in part by NIH DK39337 (M.G.S.), NIH CA45327 (W.W.S.), and the Mayo Foundation.Presented at the Society of Surgery of the Alimentary Tract, May 1995, in San Diego, California. An abstract of this work was published inGastroenterology 108:A1235, 1995.     

氨基酸腹膜透析对腹透患者营养状况的影响

目的:观察氨基酸腹膜透析液对腹膜透析患者营养状况的影响.方法:24例不卧床腹膜透析(CAPD)患者随机进入治疗组、对照组.治疗组每日使用1次0.491%的氨基酸透析液,留腹4～7h,每3个月随访1次,观察期为6个月.研究期间观察的营养观察指标包括白蛋白、标准化蛋白氮呈现率(normalized protein equivalentofnitrogen appearance,nPNA)、瘦体重百分比(%lean body mass,%LBM)、前臂肌围(mid-arm muscle circumference,MAMC)、三头肌皮皱厚度(triceps skinfold,TSF);同时观察了前白蛋白(prealbumin)、转铁蛋白(transferrin)的变化及透析充分性指标的改变.结果:治疗组氨基酸透析3个月后白蛋白[(38.67±6.81)g/L vs(42.44±3.60)g/L,P＜0.05]、转铁蛋白[(2.74±0.37)g/Lvs(2.93±0.16)g/L,P＜0.05]、前白蛋白[(461.38±330.29)g/Lvs(641.57±673.91)g/L,P＜0.05]呈明显上升,之后保持在稳定水平;对照组上述指标则逐步下降.治疗组nPNA、%LBM、MAMC在研究期间始终稳定,对照组则随透析时间的延长降低明显.两组患者透析充分性指标在氨基酸透析期间始终保持充分状态.结论:氨基酸腹膜透析可以预防及纠正CAPD患者营养不良的发生及发展.     

肝炎后肝硬变患者脑脊液游离氨基酸谱的研究与探索

本文对40例肝炎后肝硬变，13例正常人的脑脊液中16种氨基酸水平进行了检测，并同时检测了血清中氨基酸水平。结果表明：肝炎后肝硬变患者血浆氨基酸谱与脑脊液氨基酸谱完全不同。作者对肝炎后肝硬变患者脑脊液中氨基酸代谢的特点和失衡机理进行了探讨，并探讨了测定脑脊液中氨基酸水平的临床意义；（1）鉴别肝性昏迷和非肝性昏迷。（2）对于判定肝组织损伤程度，判定肝性脑病发生率和判定肝硬变预后较血清氨基酸敏感。（3）指导临床应用支链氨基酸的用时，用法和用量。     

Control of catalytic efficiency by a coevolving network of catalytic and noncatalytic residues

The active sites of enzymes consist of residues necessary for catalysis and structurally important noncatalytic residues that together maintain the architecture and function of the active site. Examples of evolutionary interactions between catalytic and noncatalytic residues have been difficult to define and experimentally validate due to a general intolerance of these residues to substitution. Here, using computational methods to predict coevolving residues, we identify a network of positions consisting of two catalytic metal-binding residues and two adjacent noncatalytic residues in LAGLIDADG homing endonucleases (LHEs). Distinct combinations of the four residues in the network map to distinct LHE subfamilies, with a striking distribution of the metal-binding Asp (D) and Glu (E) residues. Mutation of these four positions in three LHEs—I-LtrI, I-OnuI, and I-HjeMI—indicate that the combinations of residues tolerated are specific to each enzyme. Kinetic analyses under single-turnover conditions revealed that I-LtrI activity could be modulated over an ∼100-fold range by mutation of residues in the coevolving network. I-LtrI catalytic site variants with low activity could be rescued by compensatory mutations at adjacent noncatalytic sites that restore an optimal coevolving network and vice versa. Our results demonstrate that LHE activity is constrained by an evolutionary barrier of residues with strong context-dependent effects. Creation of optimal coevolving active-site networks is therefore an important consideration in engineering of LHEs and other enzymes.The active sites of enzymes are often the most conserved positions in a multiple sequence alignment, as purifying selection for maintenance of function constrains amino acid variation of residues that directly participate in catalysis. Noncatalytic residues, often in close proximity to catalytic residues, contribute to enzymatic function by maintaining the architecture and chemical environment of the active site. Noncatalytic residues often show sequence variation in multiple sequence alignments, yet nonpermissive substitutions at these positions will have an impact on enzymatic function by disrupting the architecture or chemical environment necessary for catalysis. Thus, catalytic and noncatalytic residues must coevolve with each other and surrounding residues to maintain active-site conformation and chemistry and to buffer against potentially deleterious mutations (1, 2). Coevolving residues within a protein family can be predicted by computational methods that use mutual information theory to identify residue covariation in a multiple sequence alignment (2–5). However, because the magnitude of covariation between positions varies with the magnitude of positional variation (4), the identification of catalytic residues as part of coevolving networks is problematic for the simple reason that catalytic residues show little sequence variation. Although others have identified putative coevolution between residues involved in catalysis in a small number of protein families (6–8), to our knowledge there are no examples demonstrating the functional consequences of coevolution between catalytic and surrounding noncatalytic positions.We examined coevolution between the catalytic and noncatalytic residues of LAGLIDADG homing endonucleases (LHEs), site-specific DNA endonucleases that are typically encoded within self-splicing introns and inteins (9). LHEs show extreme sequence variation and function as homodimers or as single-chain monomers composed of two LAGLIDADG domains that evolved by gene duplication or gene fusion events. The active site of LHEs consists of two parallel α-helices containing the class-defining LAGLIDADG amino acid motif with acidic metal-binding residues (D or E) at the bottom of each α-helix and positioned in close proximity to the DNA substrate (10–12). Outside of the LAGLIDADG α-helices, the extreme sequence variation makes it difficult to build robust alignments and infer functional information (13–15). Moreover, the monomeric and dimeric LHEs likely evolved under different functional constraints, and the phylogenetic signal and functional information for either form is diluted in alignments that include both the monomeric and dimeric LHEs. Because LHEs are currently under investigation for use as genome-editing agents (16–18), a greater understanding of their functional constraints would aid in engineering studies.Here, we take advantage of high-quality structure-guided multiple sequence alignments of single-chain LHEs to predict coevolving networks using methods based on mutual information (5, 19, 20). Strikingly, the network with the strongest predictive scores included the metal-binding catalytic residues and adjacent noncatalytic residues that lie on opposite LAGLIDADG α-helices. The coevolving network was experimentally confirmed by functional analyses showing that catalytic activity could be modulated over an ∼100-fold range by mutation of either catalytic or noncatalytic residues in the network. Our results show that maintaining integrity of coevolving networks of catalytic and noncatalytic residues is an important consideration when engineering LHEs and may also be applicable to engineering of other enzyme families.     

贵州两个不同地区周期型马来丝虫成虫氨基酸的组成研究

应用高效液相色谱对贵州省内的两个不同地区周期型马来丝虫（贵州独山株、贵州荔波株）成虫的１８种氨基酸进行了检测和分析。结果显示贵州独山株与荔波株马来丝虫均检出１７种氨基酸，两株丝虫有１６种相同氨基酸。两株氨基酸的总含量无明显差异（Ｐ＞０．０５）。但独山株有丝氨酸，缺酪氨酸，而荔波株含酪氨酸，缺丝氨酸。     

General approach to reversing ketol-acid reductoisomerase cofactor dependence from NADPH to NADH

To date, efforts to switch the cofactor specificity of oxidoreductases from nicotinamide adenine dinucleotide phosphate (NADPH) to nicotinamide adenine dinucleotide (NADH) have been made on a case-by-case basis with varying degrees of success. Here we present a straightforward recipe for altering the cofactor specificity of a class of NADPH-dependent oxidoreductases, the ketol-acid reductoisomerases (KARIs). Combining previous results for an engineered NADH-dependent variant of Escherichia coli KARI with available KARI crystal structures and a comprehensive KARI-sequence alignment, we identified key cofactor specificity determinants and used this information to construct five KARIs with reversed cofactor preference. Additional directed evolution generated two enzymes having NADH-dependent catalytic efficiencies that are greater than the wild-type enzymes with NADPH. High-resolution structures of a wild-type/variant pair reveal the molecular basis of the cofactor switch.     

五种丝虫的氨基酸与化学元素

用高效液相（ＨＰＬＣ）研究人、牛、马、驴、骡与犬体内５种丝虫１８种氨基酸，人、畜丝虫分别含有１６和１７种，缺色氨酸。家畜丝虫氨基酸含量（μｇ／ｍｇ干粉）高于马来丝虫，其酸性和碱性氨基酸（Ｐ＜０．０５）及支链族和芳香族氨基酸（Ｐ＜０．０１）含量高低差异明显并非常显著。以原子吸收分光光度计测定丝虫含Ｚｎ，Ｃｕ，Ｆｅ，Ｍｎ，Ｃｄ５种微量元素中以Ｚｎ为高（μｇ／０．１ｇ干粉），２种常量元素中Ｃａ多于Ｍｇ，这将为研究丝虫（病）的生理代谢与免疫学诊断提供参考资料。