水稻红莲型细胞质雄性不育幼穗发育过程中组织型转谷氨酰胺酶活性比较

以水稻红莲型细胞质雄性不育系粤泰A和保持系粤泰B的叶片和不同发育时期幼穗为材料,比较分析了水稻红莲型细胞质雄性不育幼穗发育过程中组织型转谷氨酰胺(tTG)酶活性变化,建立了适合水稻tTG酶活性分析的酶联免疫法（ELISA）测定反应体系。研究发现tTG酶活性受钙离子正调控,并且无论是粤泰A还是粤泰B,衰老叶片中tTG酶活性都高于新叶片,但粤泰A与粤泰B之间的差异不明显;不育系粤泰A自四分体到二核期的幼穗不同发育阶段,tTG酶活性随着花粉发育而增强,在二核期达到最高,而在保持系粤泰B中,tTG酶活性没有随发育进程发生显著变化。推断tTG酶与花粉败育过程中的细胞程序性死亡有关。     

Comparison of Tissue Transglutaminase Activity During Young Panicle Development in Honglian-type Cytoplasmic Male Sterile Rice

Rice is the most important food crop in China, the development of hybrid rice is a great achievement to improve rice yield. The basis of heterosis utlization in hybrid rice is development and use of the cytoplasmic male sterile system. Though China is lea…     

茂源链霉菌原生质体的制备与再生

【目的】探索制备高纯度茂源链霉菌原生质悬液的条件,为原生质体融合提供支持。【方法】在茂源链霉菌菌丝体一级培养不同时间(4,8,12,16,20,24,28,32,36,40,44,48h)后,称取菌丝体干质量,确定一级培养最佳时间;将一级培养的菌丝体转接培养二级菌丝体,在不同培养时间(13,14.5,16,17.5,19,20.5h)根据原生质体制备率、再生率的综合情况及菌丝体形态显微镜观察结果,确定二级菌丝体的培养时间及培养基中添加甘氨酸的最佳质量浓度(0,2,5,10,20g/L),根据原生质体制备率和再生率确定使用溶菌酶的质量浓度(2,5,8,10,20,50mg/mL)和酶解时间(1,2,3,4,5,6,7,8h),使用不同方法(500r/min离心法与双层18μm和0.45μm滤膜过滤法)分离原生质体,对原生质体分离后的损耗情况进行比较,确定分离条件。【结果】茂源链霉菌一级菌丝体的最佳培养时间为28h;二级菌丝体培养的最佳条件为:在培养基中添加5g/L甘氨酸,培养16h,溶菌酶最适质量浓度为8mg/mL,酶解3~4h;以500r/min离心分离原生质体可以获得较纯净的原生质体悬液;茂源链霉菌原生质体制备率最高达97.82%,再生率最高达9.04%,纯度最高达87.53%。【结论】得到了制备高纯度的茂源链霉菌原生质体悬液的条件。     

Inhibiting effect of polymerization and degradation of myosin heavy chain during preheating at 30°C and 50°C on the gel-forming ability of walleye pollack surimi

ABSTRACT: To confirm the contribution of polymerization and degradation of myosin heavy chain (MHC) during preheating to the gel-forming ability of fish meat paste, walleye pollack surimi paste was preheated at 30°C and 50°C prior to heating at 80°C in the presence of various inhibitors. At 30°C, ethyleneglycol bis(2-aminoethyl ether) -N,N,N ', N '-tetraacetic acid (EGTA) and ethylenediaminetetraacitic acid (EDTA) inhibited gel formation as well as the polymerization of MHC, whereas dithiothreitol (DTT) and leupeptin promoted gel formation, which was accompanied by the enhancement of MHC polymerization and decreased MHC degradation, respectively. At 50°C, leupeptin inhibited MHC degradation and improved gel strength, whereas EGTA, EDTA and DTT had no effect on MHC polymerization and degradation and did not affect gel formation. The results demonstrate that the gel strength of cooked gel (80°C) is not affected by preheating at 30°C and 50°C and does not inhibit polymerization and degradation. Results suggest that the gel strength of cooked gel is dependent on the polymerization and degradation of MHC during preheating.     

Purification of transglutaminase from scallop striated adductor muscle and NaCl-induced inactivation

SUMMARY: Tissue type transglutaminase (TGase) was purified from scallop striated adductor muscle with successive chromatographies of DE-52 cellulose, Sephacryl S-300, and Mono Q columns. The yield and purification of the enzymatic activity was 16.6% and 101.9-fold, respectively. The molecular mass of purified enzyme was estimated to be 95 kDa by sodium dodecylsulfate–polyacrylamide gel electrophoresis analysis. Scallop TGase was Ca²⁺-dependent and strongly inactivated by ρ-chloromercuribenzoic acid, N -ethylmaleimide, Cu²⁺, and Zn²⁺, meaning it belongs to the thiol group of enzymes as well as being a mammalian enzyme. When scallop TGase was incubated in 0.5 M NaCl without substrate for 2 h at 20°C and pH 7.5, enzymatic activity decreased to 14.4% of its original. However, a conformational change in the TGase molecule was not detected by either fluorescent, ultraviolet, and circular dichroism spectra analyses compared to the enzyme incubated without NaCl. In addition, the enzyme inactivated by NaCl was partially recovered by the dilution of salt concentration, which means that the NaCl-induced inactivation process is reversible to some extent. These results suggest that NaCl-induced modulation of the TGase molecule occurs via a small conformational change.     

Pilot scale production of a non-immunogenic soluble gluten by wheat flour transamidation with applications in food processing for celiac-susceptible people

Celiac disease (CD) is an immune-mediated disease triggered by wheat gluten and related prolamins. A lifelong gluten-free (GF) diet is mandatory to normalize the intestinal mucosa. We previously found that transamidation by microbial transglutaminase of gluten was effective in suppressing the gliadin-specific inflammatory response in CD patients without influencing the main technological properties of wheat flour or semolina. In this study, we produced on a pilot scale a soluble form of transamidated gluten (soluble protein fraction, spf), characterised by a high protein content (88 mg/ml), while native gluten was dramatically reduced (32 ± 2 ppm; R5-ELISA). Using HLA-DQ8 transgenic mice as a CD model, we found suppression of interferon-γ secretion in gliadin-specific CD4⁺ T cells challenged with spf-primed dendritic cells. In terms of functional properties, spf showed both solubility and emulsifying activity values within the range of commercial soluble glutens. Notably, dough prepared by mixing rice flour with spf could leaven. After baking, blended rice bread had a higher specific volume (2.9 ± 0.1) than control rice bread (2.0 ± 0.1) and acquired wheat-like sensory features. Taken together, our results highlighted the technological value of transamidated soluble gluten to improve both nutritional and sensory parameters of GF food.     

Improvement of Cold and Thermally Induced Gelation of Giant Squid (Dosidicus gigas) Surimi

Depending on the season of capture, giant squid (Dosidicus gigas) surimi processed by isoelectric precipitation presents low gel strength. Addition of microbial transglutaminase (MTGase) and application of high isostatic pressure (300 MPa) to improve physicochemical properties were assayed for purposes of making “suwari” gels and heated gels for use in restructured products which have a raw or cooked appearance. The physicochemical properties of both pressurized and unpressurized gels induced by application of 30°C/1 h improved when MTGase was added. In contrast, addition of MTGase was less effective in gels subsequently heated at 90°C/30 min after 30°C/1 h. High pressure treatment for 30 min at 300 MPa and 15°C helped to produce gels with better mechanical and water binding properties, whether treated for 30°C/1 h only or for 30°C/1 h plus 30-min heat treatment at 90°C. High pressure treatment also reduced lightness.     

低场NMR法研究微生物转谷氨酰酶对猪肉肌原纤维蛋白凝胶功能特性的影响

用低场核磁共振(NMR)研究了微生物转谷氨酰酶(MTG)对猪肉肌原纤维蛋白凝胶质子核磁共振弛豫行为的影响.研究结果显示,NMR结果拟合后得到的水有4个组分,合并为对应水的3种状态即结合水、不易流动水和自由水.和对照相比,加酶处理后显著降低了猪肉肌原纤维蛋白热诱导凝胶的自旋-自旋弛豫时间(T2).加入2 U·g-1剂量的MTG后,T22b值(对应于不易流动水)由226 ms(峰值)降低到188 ms.然而,当酶浓度增加时,T22b没有进一步降低.对衰减NMR信号拟合结果进行主成分分析显示,前两个主成分可以解释变异方差的80%,并且加酶和对照在样品评分图上显著分开.加入MTG后,凝胶中水质子的移动性显著下降.     

鳙内源性转谷氨酰胺酶特性的研究

文章研究了温度、pH、二硫苏糖醇（DTT）以及不同的金属离子和抑制剂对鳙（Hypophthalmichthys nobilis）鱼肉中内源性转谷氨酰胺酶（transglutaminase,TGase）活性的影响。结果表明,鳙内源性TGase的最适温度在37～45℃之间,最适pH为6.5～7.5,适量的钙离子（Ca^2＋）（0～2.2 mmol·L^-1）可以激发酶活性,低浓度（0～0.2mmol·L^-1）的DTT可以提高酶活性,但高浓度的钠离子（Na^＋）和钾离子（K^＋）会降低酶活性,镁离子（Mg^2＋）、铜离子（Cu^2＋）、锌离子（Zn^2＋）、钡离子（Ba^2＋）、铁离子（Fe^3＋）、亚铁离子（Fe^2＋）、铅离子（Pb^2＋）和乙二胺四乙酸（EDTA）也会抑制酶的活性。     

Microbial transglutaminases generate T cell stimulatory epitopes involved in celiac disease

Celiac disease (CD) is a permanent intolerance to gluten. In CD patients, gluten peptides cause an inflammation in the small intestine leading to tissue damage. Tissue transglutaminase (tTG) is an enzyme involved in the repair of damaged tissue by crosslinking of extracellular matrix proteins. Under certain conditions, tTG can deamidate glutamine into glutamic acid. Compared to native gluten, deamidated gluten elicits a more powerful inflammatory response. To improve the quality and texture of food products microbial transglutaminases (mTG) are used in the food industry.