重组人胸腺肽α1在大肠杆菌中的表达纯化和鉴定

用PCR将化学合成的人胸腺肽α1基因扩增并克隆到pMD18-T载体中,经过KpnI／SacI酶切、连接将胸腺肽基因插入到表达载体pET32b中硫氧还蛋白下游．将重组质粒转化至表达宿主BL21(DE3)中,经IPTG诱导,Zn—Sepharose亲和层析纯化以及复性处理,从每升菌液中可获得35mg硫氧还蛋白-胸腺肽α1融合蛋白．经凝血因子Xa酶切、Zn—Sepharose亲和层析以及反相高效液相色谱纯化获得3．8mg重组人胸腺肽．小鼠胸腺细胞凋亡实验表明,重组人胸腺肽能显地抑制地赛米松诱导的小鼠胸腺细胞的凋亡,且该抑制作用与重组人胸腺肽的浓度呈剂量依赖关系．     

Involvement of penaeidins in defense reactions of the shrimp Litopenaeus stylirostris to a pathogenic vibrio

The present study reports for the first time the involvement of an antimicrobial peptide in the defense reactions of a shrimp infected by a pathogenic Vibrio, Vibrio penaeicida. New members of the penaeidin family were characterized in the shrimp Litopenaeus stylirostris by RT-PCR and RACE-PCR from hemocyte total RNAs, and by mass spectrometry detection and immunolocalization of mature peptides in shrimp hemocytes. In infected shrimps, bacteria and penaeidin distribution colocalized in the gills and the lymphoid organ that represented the main infected sites. Moreover, the shrimp immune response to infection involved massive hemocyte recruitment to infection sites where released penaeidin may participate in the isolation and elimination of the bacteria, We show that the ability of the shrimps to circumvent shrimp infections is closely related to a recovery phase based on the hematopoietic process.Received 25 November 2003; received after revision 8 January 2004; accepted 21 January 2004     

Hemokinins and endokinins

The mammalian tachykinins are a family of peptides that, until recently, has included substance P (SP), neurokinin A and neurokinin B. Since, the discovery of a third preprotachykinin gene (TAC4), the number of tachykinins has more than doubled to reveal several species-divergent peptides. This group includes hemokinin-1 (HK-1) in mouse and rat, endokinin-1 (EK-1) in rabbit, and EKA, EKB, human HK-1 (hHK-1) and hHK(4–11) in humans. Each exhibits a remarkable selectivity and potency for the tachykinin NK₁ receptor similar to SP. Their peripheral expression has led to the proposal that they are the endogenous peripheral SP-like endocrine/paracrine agonists where SP is not expressed. Moreover, their strong cross-reactivity with a specific SP antibody leads us to question many of the proposed locations and roles of SP in the periphery. Additionally, three orphan tachykinin gene-related peptides are identified on TAC4, in rabbit, EK-2 and in humans, EKC and EKD.Received 25 January 2004; received after revision 18 February 2004; accepted 27 February 2004     

4-Hydroxynonenal-modified amyloid-beta peptide inhibits the proteasome: possible importance in Alzheimer's disease

The amyloid β-peptide (Aβ) is a 4-kDa species derived from the amyloid precursor protein, which accumulates in the brains of patients with Alzheimer’s disease. Although we lack full understanding of the etiology and pathogenesis of selective neuron death, considerable data do imply roles for both the toxic Aβ and increased oxidative stress. Another significant observation is the accumulation of abnormal, ubiquitin-conjugated proteins in affected neurons, suggesting dysfunction of the proteasome proteolytic system in these cells. Recent reports have indicated that Aβ can bind and inhibit the proteasome, the major cytoslic protease for degrading damaged and ubiquitin-conjugated proteins. Earlier results from our laboratory showed that moderately oxidized proteins are preferentially recognized and degraded by the proteasome; however, severely oxidized proteins cannot be easily degraded and, instead, inhibit the proteasome. We hypothesized that oxidatively modified Aβ might have a stronger (or weaker) inhibitory effect on the proteasome than does native Aβ. We therefore also investigated the proteasome inhibitory action of Aβ _1–40 (a peptide comprising the first 40 residues of Aβ) modified by the intracellular oxidant hydrogen peroxide, and by the lipid peroxidation product 4-hydroxynonenal (HNE). H₂O₂ modification of Aβ _1–40 generates a progressively poorer inhibitor of the purified human 20S proteasome. In contrast, HNE modification of Aβ _1–40 generates a progressively more selective and efficient inhibitor of the degradation of fluorogenic peptides and oxidized protein substrates by human 20S proteasome. This interaction may contribute to certain pathological manifestations of Alzheimer’s disease Received 26 September 2000; accepted 26 September 2000     

动物抗菌肽的研究及应用

抗菌肽是具有抗菌活性的一类短肽,广泛存在于生物界.研究表明,除了具有广谱抗菌活性外,抗菌肽还具有抗病毒、抗真菌、抗寄生虫及抗肿瘤等生物活性.基于对抗菌肽的分类与特性、结构与活性、作用机理等方面进行系统的分析,综述了抗菌肽研究进展及应用前景.     

超滤分离纯化麦胚蛋白酶解物的研究

采用超滤法从麦胚蛋白酶解物中初步分离纯化抗氧化活性肽,考察了操作压力、操作温度及料液浓度等工艺参数对超滤过程中膜通量的影响,并对超滤前后麦胚蛋白酶解物的相对分子质量分布、氨基酸组成及抗氧化活性进行了比较.结果表明:采用截留相对分子质量3000的聚砜膜对麦胚蛋白酶解物进行超滤分离的最适工艺条件为操作压力0.30 MPa、操作温度20 ℃、麦胚蛋白酶解物浓度35 g/L;超滤有效地除去了料液中相对分子质量较大的组分,麦胚蛋白酶解物中相对分子质量1 000～500及500～130的组分分别为23.81%、48.63%,其抗氧化活性得到提高,清除O2-·的IC50由超滤前的1.64 mg·mL-1降至超滤后的1.29 mg·mL-1.图5,表3,参13.     

L-抗坏血酸-6-对羟基苯甲酸酯的合成及抗氧化活性研究

采用直接酯化法,合成了L-抗坏血酸-6-对羟基苯甲酸酯(APHB),并从清除羟自由基、超氧阴离子自由基、DPPH自由基能力及还原能力测试4个方面研究了它的体外抗氧化活性。结果表明,L-抗坏血酸-6-对羟基苯甲酸酯对上述3种自由基均有较好的清除效果,还原能力也较强,其抗氧化性能总体上与VC相当,优于市面上常用的抗氧化剂TBHQ,是一种有潜力的VC衍生物。     

Study on peptide-peptide interaction using high-performance affinity chromatography and quartz crystal microbalance biosensor

The specific interaction between sense and antisense peptides was studied by high-performance affinity chromatography （HPAC） and quartz crystal microbalance （QCM） biosensor. Fragment 1-14 of human interferon-β （hlFN-βwas chosen as sense peptide and its three antisense peptides （AS-IFN 1, AS-IFN 2, and AS-IFN 3） were designed according to the degeneracy of genetic codes. The affinity column was prepared with sense peptide as ligand and the affinity chromatographic behavior was evaluated. Glu-substituted antisense peptide （AS-IFN 3） showed the strongest binding to immobilized sense peptide at pH 7.5. A quartz crystal microbalance-flow injection analysis （QCM-FIA） system was introduced to investigate the recognition process in real-time. The equilibrium dissociation constants between sense peptide and AS-IFN 1, AS-IFN 2 and AS-IFN 3 measured 2.08×10^-4, 1.31×10^-4 and 2.22×10^-5 mol/L, respectively. The mechanism study indicated that the specific recognition between sense peptide and AS-IFN 3 was due to sequence-dependent and multi-modal affinity interaction.     

Transport of a low molecular weight extracellular esterase into membrane vesicles ofCandida lipolytica

Summary The low mol. wt extracellular esterase ofCandida lipolytica is actively transported into membrane vesicles. In the absence of metabolic energy, a proton gradient can drive the transport process. The transport system does not accumulate the enzyme at peak levels due to the presence of a leak pathway.     

Chemical composition and antioxidant activity of essential oil from Magnolia grandiflora L. seed

The chemical composition and antioxidant activity of Magnolia grandiflora L.seed essential oil were examined.By combining gas chromatography and mass spectrometry (GC-MS),16 types of compounds were identified,including 40.91% oxygen-containing terpene derivatives,19.51% terpenes,13.97% esters,13.05% nitrogenous compounds,and 3.64% acids.Its capability of removing hydroxyl free radical was lower than that of V c,but it was closer to V c with increasing concentration.In cell experiment,all essential oil concentration (6.9,13.8,27.7,55.5,111.0,and 222.0 g/mL) groups increased cell proliferation by 26% or more (26.0%,38.8%,44.8%,40.2%,35.8%,and 30.4%,respectively).In rat experiment,the SD rats group showed an at least 38% rise (38.62%,61.68%,and 62.86%,respectively) in glutathione-peroxisome (GSH-Px)activity,which is more than 24% reduction (29.56%,24.30%,and 36.93%) in malondialdehyde (MDA) content,and no significant difference in SOD activity was found.The results show that the essential oil had strong antioxidant activity,without dose effect within the concentration range.