大肠杆菌细胞壁肽聚糖的化学修饰及荧光标记

【目的】发展一种活细菌细胞壁荧光标记方法,为后期研究细菌肽聚糖的生物合成和代谢规律以及其与细菌感染致病的关系提供新的工具。【方法】对细菌肽聚糖的生物合成前体N-乙酰葡萄糖胺-1-磷酸(GlcNAc-1-P)进行化学修饰,设计并合成含有叠氮基的GlcNAc-1-P类似物(化合物5:Ac3GlcNAz-1-P)。将该类似物与细菌共同孵育,使其作为探针进入细菌肽聚糖天然合成途径。之后提取并酶解肽聚糖组分,用红外光谱(FTIR)和液质联用(LC/MS)检测探针是否通过代谢进入细菌肽聚糖结构中。同时用外源荧光素对代谢掺入细菌肽聚糖中的探针进行染色。在激光共聚焦显微镜下观察对活细菌的荧光标记效果。【结果】通过四步有机合成反应,以79%的总收率成功获得了化合物5。将大肠杆菌(Escherichia coli BL21)作为模式菌株与化合物5共孵育后,其肽聚糖组分的LC/MS和FTIR分析结果均显示探针可以被细菌利用并被代谢掺入到肽聚糖结构中。激光共聚焦显微镜观察结果显示,荧光素可以高效标记表面携带有生物正交探针的大肠杆菌。【结论】设计合成了一种新型探针,可用于活细菌成像,为深入研究细菌肽聚糖的生物学功能及其与细菌感染致病的关系提供了一种简便的方法。     

从效应蛋白视角看革兰氏阴性细菌VI型蛋白分泌系统底物转运机理

蛋白质分泌系统是细菌与外界交流的重要工具。革兰氏阴性细菌的Ⅵ型蛋白分泌系统(T6SS)可以转运分泌蛋白至细菌和真核细胞内,在菌间竞争中发挥重要作用,是细菌的一种重要的生存适应性武器。分泌蛋白主要包括起到运载作用的结构蛋白和有细胞毒性的效应蛋白这两类。本文主要从效应蛋白的视角讨论T6SS如何识别并转运效应蛋白的作用机理,回顾了以VgrG和PAAR为端部载体蛋白的转运途径、依赖端部运输的效应蛋白、T6SS伴侣蛋白等重要发现的背景和过程,并综述了T6SS分泌途径的新进展。     

从效应蛋白视角看革兰氏阴性细菌Ⅵ型蛋白分泌系统底物转运机理

蛋白质分泌系统是细菌与外界交流的重要工具。革兰氏阴性细菌的Ⅵ型蛋白分泌系统(T6SS)可以转运分泌蛋白至细菌和真核细胞内,在菌间竞争中发挥重要作用,是细菌的一种重要的生存适应性武器。分泌蛋白主要包括起到运载作用的结构蛋白和有细胞毒性的效应蛋白这两类。本文主要从效应蛋白的视角讨论T6SS如何识别并转运效应蛋白的作用机理,回顾了以VgrG和PAAR为端部载体蛋白的转运途径、依赖端部运输的效应蛋白、T6SS伴侣蛋白等重要发现的背景和过程,并综述了T6SS分泌途径的新进展。     

双歧杆菌肽聚糖结构及分子量的分析

从双歧杆菌细胞壁中分离纯化肽聚糖,研究其化学成分、结构和分子量分析。经氨基酸、多糖和蛋白质含量分析和肽聚糖的溶解实验鉴定,所提取物质的主要成分为肽聚糖;经核磁共振和红外光谱分析,所提取肽聚糖的结构与已知的肽聚糖的结构相吻合,其糖链结构为由D型吡喃糖N-乙酰葡糖胺和N-乙酰胞壁酸以β-1,4-糖苷键连接而组成;经SDS-PAGE和多种染色方法结合,分析经溶菌酶水解后的肽聚糖,其分子量的分布呈弥散状,范围在97.6kD到14.4kD之间。     

人源溶菌酶结构与功能的研究进展

人溶菌酶是一类人体内天然存在的能够溶解细菌细胞壁的碱性蛋白的总称。其作用特征是能够裂解肽聚糖中的N-乙酰氨基葡萄糖与N-乙酰氨基甲酸之间的β-(1,4)-糖苷键。人溶菌酶具有抗菌、抗炎、抗病毒和增强免疫力等多种特性,因此在国内外市场上应用广泛。本文就人溶菌酶的结构特点、表达部位、功能表达以及应用情况进行综述。     

鼠伤寒沙门氏菌Ⅵ型分泌系统相关基因敲除及其功能特性

Ⅵ型分泌系统(T6SS)是大多数革兰氏阴性细菌中都存在的一种重要的分泌系统,能介导细菌与细菌之间以及细菌和宿主细胞之间的相互作用,溶血素共调节蛋白(Hcp)和缬氨酸甘氨酸重复蛋白G(VgrG)是组成T6SS穿刺装置的重要组分。但鼠伤寒沙门氏菌Ⅵ型分泌系统的Hcp与VgrG在该菌入侵宿主细胞及抗吞噬过程中发挥的作用尚不十分清楚。【目的】本研究旨在利用基因敲除技术构建的鼠伤寒沙门氏菌hcp及vgrg基因缺失株体外接种真核上皮细胞和巨噬细胞,并以其亲本株作为对照,以研究Hcp及VgrG在该菌粘附、侵入上皮细胞及抗吞噬过程中所发挥的作用。【方法】通过优化Red同源重组系统操作过程中各个条件,建立一套快速敲除鼠伤寒沙门氏菌Ⅵ型分泌系统相关基因的操作系统,成功构建鼠伤寒沙门氏菌CVCC541的hcp及vgrg单基因缺失株、双基因缺失株及三基因缺失株,并用Hela细胞接种试验和菌落计数试验,评估不同菌株的粘附和侵袭能力;用小鼠巨噬细胞RAW 264.7接种试验,评估不同菌株的抗吞噬能力。【结果】与亲本株CVCC541粘附侵袭Hela细胞相比,基因缺失株CVCC541Δvgrg、CVCC541Δhcp2Δvgrg和CVCC541Δhcp1Δhcp2Δhcp3的粘附率分别为17.17%±2.1%、14.73%±2.5%和82%±3.7%;CVCC541Δvgrg、CVCC541Δhcp2Δvgrg和CVCC541Δhcp1Δhcp2Δhcp3的侵袭率分别为7.05%±1.05%、6.21%±1.35%和87%±3.25%;与亲本株CVCC541在小鼠巨噬细胞RAW 264.7中的存活相比,基因缺失株CVCC541Δvgrg、CVCC541Δhcp2Δvgrg和CVCC541Δhcp1Δhcp2Δhcp3的存活率分别为15.67%±2.9%、14.47%±1.87%和56.12%±3.48%。【结论】鼠伤寒沙门氏菌Ⅵ型分泌系统VgrG和Hcp对该菌入侵细胞和抗吞噬方面具有重要作用,该研究为鼠伤寒沙门氏菌通过六型分泌系统与宿主细胞相互作用的机制研究奠定了基础。     

镧和铈对蜡状芽孢杆菌抗性、生长及胞内核酸的影响

【目的】研究稀土元素对细菌生理效应的影响及机理。【方法】用抑菌圈法和光电比浊法测定了硝酸镧和硝酸铈对蜡状芽孢杆菌抗性及生长的影响, 并用傅立叶变换红外光谱和荧光分光光度法研究了细菌细胞壁及胞内DNA的结构。【结果】40?50 mg/L硝酸镧增强了细菌对9%石炭酸, 0.135%升汞, 8×105 U/mL青霉素的抗性; 40?50 mg/L硝酸铈增强了细菌对9%石炭酸, 0.135%升汞, 8×105 U/mL青霉素和75%次氯酸钠的抗性; 稀土元素会加快蜡状芽孢杆菌生长和繁殖速度, 同时高浓度稀土元素的促进作用要比低浓度时更明显, 表明稀土元素可以刺激细菌繁殖; 红外光谱表明稀土元素会影响细菌细胞壁肽聚糖结构, 荧光分光光度法结果显示稀土元素会影响细菌胞内DNA的结构。【结论】细菌抗性的改变可能与细胞壁肽聚糖结构受到影响有关; 稀土元素对DNA的影响可能是抗性和生长受到影响的根本原因。     

昆虫肽聚糖识别蛋白研究进展

在脊椎动物和非脊椎动物中,识别非己是天生免疫反应中的第一步。肽聚糖是细菌细胞壁的必需成分,属于进化上保守的微生物表面病原相关分子模式(pathogen-associated molecular pattern, PAMP),可以被模式识别蛋白(pattern recognition proteins, PRRs)如肽聚糖识别蛋白(peptidoglycan recognition proteins, PGRPs)识别。 在昆虫的天生免疫系统中,有些PGRPs能够利用细菌独有的肽聚糖识别入侵细菌,并将细菌入侵信号传递给下游的抗菌肽(antimicrobial peptide, AMP)合成途径,启动抗菌肽基因的转录及合成;PGRPs对肽聚糖的识别也会启动酚氧化酶原途径的激活,引起黑化反应。有些具有酰胺酶活性的PGRPs可以促进吞噬作用;有些可以抑制抗菌肽合成以减弱过度免疫反应带来的损伤。还有一些PGRPs作为效应因子直接作用于细菌将细菌杀死。本文主要从昆虫PGRPs作为识别受体(recognition receptor)、调节子(regulator)和效应因子(effector) 3个方面进行了综述,并分析了目前PGRPs研究中仍不清楚的问题和未来研究的方向。     

鼠伤寒沙门氏菌Ⅲ型分泌系统抗感染类抑制剂的筛选

【背景】肠道沙门氏菌(Salmonella enterica)是一种常见的食源性肠道致病菌,可以感染人畜并引发食物中毒、伤寒等疾病。近年来因抗生素滥用导致肠道沙门氏菌耐药性问题日益严峻,迫切需要开发新型抗感染药物。肠道沙门氏菌致病的关键在于与宿主细胞接触后可以通过Ⅲ型分泌系统(type Ⅲ secretion system, T3SS)向宿主细胞内注射效应蛋白,进而调控宿主细胞囊泡运输和免疫应答等生理活动,以方便其高效侵染宿主细胞。T3SS是一类由超过20种蛋白质组成、高度复杂的跨膜分子机器,是革兰氏阴性病原菌中普遍存在的一类蛋白质运输系统和毒力系统。在不同病原菌中,其结构与功能非常保守。位于T3SS核心跨膜区的SctV家族蛋白是T3SS中最保守的组分之一,参与T3SS能量供应和效应蛋白的分泌过程,SctV蛋白的关键氨基酸突变失活后会导致鼠伤寒沙门氏菌丧失对宿主的入侵能力。【目的】以沙门氏菌SctV家族蛋白为靶点,尝试通过虚拟筛选技术筛选与SctV胞内区相互作用的抗感染类T3SS抑制剂。【方法】结合体外相互作用分析、细菌生长曲线实验、细菌分泌实验和细胞侵染实验等对候选分子进行抑制效果的...     

枯草芽孢杆菌SS6N-酰基高丝氨酸内酯酶基因的克隆表达及其酶学特性

摘要:【目的】从一株具有细菌群体感应(Quorum Sensing,QS)信号分子淬灭活性的枯草芽孢杆菌(Bacillus subtilis) SS6中扩增N-酰基高丝氨酸内酯酶(N-acylhomoserine lactonase,AiiA)基因aiiASS6并异源表达,研究此信号降解酶的酶学特性。【方法】设计特异性引物,从B.subtilis SS6中克隆N-酰基高丝氨酸内酯酶基因aiiASS6,测序并进行生物信息学分析;将此基因克隆到表达载体pET28(a),构建重组菌株并提纯目的蛋白AiiASS6;然后用高效液相色谱(high performance liquid chromatography,HPLC)分析目的蛋白AiiASS6降解QS信号分子N-(3-Oxooctanoyl)-L-homoserine lactone (OOHL)的酶学特性。【结果】克隆得到基因片段,命名为 aiiASS6 (GenBank: KP125494),其编码一条含有297氨基酸残基的多肽,用pET28(a)成功构建重组质粒pET28-aiiASS6。生物信息学分析表明,AiiASS6的氨基酸序列含有N-酰基高丝氨酸内酯酶典型的“HXHXDH”基序和194 位的Tyr残基。在Escherichia coli BL21(DE3)中异源表达AiiASS6,用Ni柱纯化后,AiiASS6含量达2.76 mg/mL。HPLC检测结果表明AiiASS6对OOHL具有很强的催化活性及耐热性,Km和Vmax分别为0.998 mmol/L和22.3 U/mg,最适pH为7.6,最适温度范围为50－90℃;此酶在4℃保存3个月后其残余活性仍达到86%,表现出较强的稳定性。【结论】从B．subtilis SS6中获得的QS淬灭酶AiiASS6表现出降解QS信号分子的高活性,其酶学特性表明它具有作为微生物制剂防控植物或水产养殖中基于QS调控的病原菌毒力的应用潜力。     

Unusual composition of peptidoglycan in Bordetella pertussis

The composition of the peptidoglycan of Bordetella pertussis and the nature of its turnover products was determined by a new combination of analytical techniques: high performance liquid chromatography of an enzymatic peptidoglycan hydrolysate and fast atom bombardment mass spectrometry and fast atom bombardment collision-activated dissociation tandem mass spectrometry. Sixteen major components of the peptidoglycan were purified, and assignment of complete or partial chemical structures was achieved for nine and seven species, respectively. At this level of resolution, a previously unrecognized heterogeneity of monomeric (five new species; nine total) and dimeric species (five new species; five total) was detected. No species containing diaminopimelyl-diaminopimelic acid cross-links or lysyl-arginine substitutions were found. Previous estimates of total cross-linkage and average chain length were revised downward to 32% and 21 disaccharide residues, respectively. Detection of a chemically novel species, a disaccharide octapeptide monomer, in both the peptidoglycan hydrolysate and culture supernatant fluid, suggests that an N-acetyl-muramyl-L-alanine amidase acts on the intact peptidoglycan of Bordetella and participates in cell wall turnover. Five peptidoglycan turnover products were identified in the supernatant fluid of late logarithmic phase cultures, including the 1,6-anhydro monomeric species known as tracheal cytotoxin. Peptidoglycan turnover was detected at a low rate of approximately 10%/generation, a value sufficient to account for the generation of all tracheal cytotoxin found in culture supernatant fluids.     

The metabolic fate of 14C-labeled immunoadjuvant peptidoglycan monomer. II. In vitro studies

Peptidoglycan monomer (GlcNAc-MurNAc-L-Ala-D-isoglutamine-meso-diaminopimelic acid-D-Ala-D-Ala), labeled with 14C both in the disaccharide and pentapeptide portions, was incubated with slices of mouse liver, kidney or spleen as well as with mouse and human blood, blood cells plasma and serum. Peptidoglycan monomer was isolated unchanged after incubations with mouse organs and blood cells. However, upon incubation with mouse or human blood, 10-50% of the peptidoglycan monomer underwent hydrolysis to the corresponding disaccharide and pentapeptide. After incubations with plasma and serum more than 90% of the [14C]peptidoglycan monomer was metabolized: about 50% of the administered radioactive dose was recovered in the disaccharide unit and about 35% in the pentapeptide part. These results suggest that in blood, plasma and serum of mouse and man, an N-acetylmuramoyl-L-alanine amidase (mucopeptide amidohydrolase, EC 3.5.1.28) exists which splits the amide bond between the lactyl carboxyl group of the muramyl residue and the amino group of the peptide moiety in the peptidoglycan molecule.     

Structure of Bordetella pertussis peptidoglycan.

Bordetella pertussis Tohama phases I and III were grown to the late-exponential phase in liquid medium containing [3H]diaminopimelic acid and treated by a hot (96 degrees C) sodium dodecyl sulfate extraction procedure. Washed sodium dodecyl sulfate-insoluble residue from phases I and III consisted of complexes containing protein (ca. 40%) and peptidoglycan (60%). Subsequent treatment with proteinase K yielded purified peptidoglycan which contained N-acetylglucosamine, N-acetylmuramic acid, alanine, glutamic acid, and diaminopimelic acid in molar ratios of 1:1:2:1:1 and less than 2% protein. Radiochemical analyses indicated that 3H added in diaminopimelic acid was present in peptidoglycan-protein complexes and purified peptidoglycan as diaminopimelic acid exclusively and that pertussis peptidoglycan was not O acetylated, consistent with it being degraded completely by hen egg white lysozyme. Muramidase-derived disaccharide peptide monomers and peptide-cross-linked dimers and higher oligomers were isolated by molecular-sieve chromatography; from the distribution of these peptidoglycan fragments, the extent of peptide cross-linking of both phase I and III peptidoglycan was calculated to be ca. 48%. Unambiguous determination of the structure of muramidase-derived peptidoglycan fragments by fast atom bombardment-mass spectrometry and tandem mass spectrometry indicated that the pertussis peptidoglycan monomer fraction was surprisingly homogeneous, consisting of greater than 95% N-acetylglucosaminyl-N-acetylmuramyl-alanyl-glutamyl-diaminopimelyl++ +-alanine.     

In vitro metabolism of 2,2'-diaminopimelic acid from gram-positive and gram-negative bacterial cells by ruminal protozoa and bacteria.

Bacillus megaterium GW1 and Escherichia coli W7-M5 were specifically radiolabeled with 2,2'-diamino[G-3H]pimelic acid [( 3H]DAP) as models of gram-positive and gram-negative bacteria, respectively. These radiolabeled bacterial mutants were incubated alone (control) and with mixed ruminal bacteria or protozoa, and the metabolic processes, rates, and patterns of radiolabeled products released from them were studied. Control incubations revealed an inherent difference between the two substrates; gram-positive supernatants consistently contained 5% radioactivity, whereas even at 0 h, those from the gram-negative mutant released 22%. Incubations with ruminal microorganisms showed that the two mutants were metabolized differently and that protozoa were the major effectors of their metabolism. Protozoa exhibited differential rates of engulfment (150 B. megaterium GW1 and 4,290 E. coli W7-M5 organisms per protozoan per h), and they extensively degraded [3H]DAP-labeled B. megaterium GW1 at rates up to nine times greater than those of ruminal bacteria. By contrast, [3H]DAP-labeled E. coli W7-M5 degradation by either ruminal bacteria or ruminal protozoa was more limited. These fundamental differences in the metabolism of the two mutants, especially by ruminal protozoa, were reflected in the patterns and rates of radiolabeled metabolites produced; many were rapidly released from [3H]DAP-labeled B. megaterium GW1, whereas few were slowly released from [3H]DAP-labeled E. coli W7-M5. Most radiolabeled products derived from [3H]DAP-labeled B. megaterium GW1 were peptides of bacterial peptidoglycan origin. The ruminal metabolism of DAP-containing gram-positive and gram-negative bacteria, even with the same peptidoglycan chemotype, is thus likely to be profoundly different.(ABSTRACT TRUNCATED AT 250 WORDS)     

In vitro metabolism of 2,2'-diaminopimelic acid from gram-positive and gram-negative bacterial cells by ruminal protozoa and bacteria

Bacillus megaterium GW1 and Escherichia coli W7-M5 were specifically radiolabeled with 2,2'-diamino[G-3H]pimelic acid [( 3H]DAP) as models of gram-positive and gram-negative bacteria, respectively. These radiolabeled bacterial mutants were incubated alone (control) and with mixed ruminal bacteria or protozoa, and the metabolic processes, rates, and patterns of radiolabeled products released from them were studied. Control incubations revealed an inherent difference between the two substrates; gram-positive supernatants consistently contained 5% radioactivity, whereas even at 0 h, those from the gram-negative mutant released 22%. Incubations with ruminal microorganisms showed that the two mutants were metabolized differently and that protozoa were the major effectors of their metabolism. Protozoa exhibited differential rates of engulfment (150 B. megaterium GW1 and 4,290 E. coli W7-M5 organisms per protozoan per h), and they extensively degraded [3H]DAP-labeled B. megaterium GW1 at rates up to nine times greater than those of ruminal bacteria. By contrast, [3H]DAP-labeled E. coli W7-M5 degradation by either ruminal bacteria or ruminal protozoa was more limited. These fundamental differences in the metabolism of the two mutants, especially by ruminal protozoa, were reflected in the patterns and rates of radiolabeled metabolites produced; many were rapidly released from [3H]DAP-labeled B. megaterium GW1, whereas few were slowly released from [3H]DAP-labeled E. coli W7-M5. Most radiolabeled products derived from [3H]DAP-labeled B. megaterium GW1 were peptides of bacterial peptidoglycan origin. The ruminal metabolism of DAP-containing gram-positive and gram-negative bacteria, even with the same peptidoglycan chemotype, is thus likely to be profoundly different.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization of Staphylococcus aureus cell wall glycan strands, evidence for a new beta-N-acetylglucosaminidase activity

Using sequential digestion with the glycyl-glycine endopeptidase lysostaphin followed by the pneumococcal N-acetylmuramyl-L-alanine amidase (amidase), the glycan strands of the peptidoglycan of Staphylococcus aureus were purified and analyzed by a combination of reverse-phase-high pressure liquid chromatography (HPLC) and mass spectrometry. Reverse-phase-HPLC resolved the glycan strands to a family of major peaks, which represented oligosaccharides composed of repeating disaccharide units (N-acetylglucosamine-[beta-1, 4]-N-acetylmuramic acid) with different degrees of polymerization and terminating with N-acetylmuramic acid residues at the reducing ends. The method allowed separation of strands up to 23-26 disaccharide units with a predominant length between 3 and 10 and an average degree of polymerization of approximately 6. Glycan strands with a higher degree of polymerization (>26 disaccharide units) represented 10-15% of the total UV absorbing glycan material. A unique feature of the staphylococcal glycan strands was the presence of minor satellite peaks that were present throughout the HPLC elution profile eluting either just prior or shortly after the major oligosaccharide peaks. A number of observations including mass spectrometric analysis suggest that the satellites are the products of an N-acetylglucosaminidase activity that differs from the atl gene product and that appears to be involved with modification of the glycan strand structure.     

家蚕抗菌肽CM4对大肠杆菌超微结构影响的研究（简报）

The effect of antibacterial peptide CM4 of Bombyx mori against E. coli K12 was investigated using scanning electron microscopy(SEM) and transmission electron microscopy (TEM). The ultrastructural changes of E. coli K12 were observed by the challenge of the purified antibacterial peptide CM4. The results showed that the antibacterial peptide caused a series of pathological changes on E. coli. SEM and TEM revealed aggregates of bacteria and SEM revealed wrinkled bacterial surfaces in the early stage. Thereafter, plasmolysis was observed with irregular holes appearing in the two ends of bacteria and the cytoplasmic contents of the cells leaking out. Finally, bacteria became empty vesicles and disintegrated into small fragments subsequently. Comparatively, the bacterial membrane was normal and the bacterial structure remained intact in the control group.     

家蚕抗菌肽CM4对大肠杆菌超微结构影响的研究*(简报)

The effect of antibacterial peptide CM4 of Bombyx mori against E. coll K12 was investigated using scanning electron microscopy(SEM) and transmission electron microscopy (TEM). The ultrastructural changes of E. coli K12 were observed by the challenge of the purified antibacterial peptide CM4. The results showed that the antibacterial peptide caused a series of pathological changes on E. coli. SEM and TEM revealed aggregates of bacteria and SEM revealed wrin-kled bacterial surfaces in the early stage. Thereafter, plasmolysis was observed with irregular holes appearing in the two ends of bacteria and the cytoplasmic contents of the cells leaking out. Finally, bacteria became empty vesicles and disintegrated into small fragments subsequently. Comparatively, the bacterial membrane was normal and the bacterial structure remained intact in the control group.     

Promotion of sleep by gonococcal peptidoglycan fragments. Structural requirements for the somnogenic activity

We exploited an extensive inventory of gonococcal peptidoglycan (PG) fragments to define the essential structural determinants of PG-mediated sleep-promoting activity in a rabbit sleep model. PG fragments, purified using reverse phase HPLC and structurally defined by fast atom bombardment mass-spectrometry, were administered intracerebroventricularly and the duration of specific sleep stages was determined electroencephalographically. Of the compounds tested, the principal naturally occurring sleep factor isolated from sleep-deprived animals (N-acetylglucosaminyl-[NAG]-1,6-anhydro-N-acetylmuramyl[anh.NAM]-alanyl-glutamyl-diaminopimelyl-alanine), the structurally identical PG monomer derived from gonococci, and individual analogs of the gonoccocal compound which lacked the NAG residue or contained an additional alanine at the C-terminus possessed maximal potency; as little as 1 pmol of these anh.NAM-containing monomers induced excess slow wave sleep (p < .05). In fact, each of five different anh.NAM-containing disaccharide peptides tested was somnogenic at 10 pmol or less, but none of a matched set of analogous PG monomers, differing only in replacement of anh.NAM by a hydrated NAM residue, was somnogenic at this dose. Together, these data suggested that the anh.NAM end, but not the NAG moiety, is a crucial structural determinant of gonococcal PG-mediated somnogenic activity. The somnogenic activity of anh.NAM-containing fragments was also modulated (albeit to a lesser extent) by the length and composition of the peptide side chain. On a much broader basis, the data also help raise the intriguing hypothesis that bacterial products may serve as natural regulators of nervous system function in higher animals.     

Autoradiographic analysis of diaminopimelic acid incorporation in filamentous cells of Escherichia coli: repression of peptidoglycan synthesis around the nucleoid.

Peptidoglycan synthesis rate in nonconstricting filaments of Escherichia coli dnaX(Ts) has been studied by autoradiography of incorporated [3H]diaminopimelic acid. Analysis of autoradiograms of whole cells and sacculi showed that peptidoglycan is synthesized at a reduced rate in the nucleoid-containing parts of these filaments. The lower rate of peptidoglycan synthesis in the cell center coincides with a higher local rate of protein synthesis. DNA-less cell formation in dnaX(Ts), dnaX(Ts) sfiA, and the minB minicell-forming mutant is accompanied by a local increase in peptidoglycan synthesis at the constriction site.