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本试验旨在研究绵羊瘤胃细菌、原虫蛋白质分解代谢相关酶活力及谷氨酸脱氢酶体系的米氏常数(Km)值,为解释绵羊瘤胃细菌、原虫蛋白质分解代谢特征提供酶学依据。选用6只1岁左右安装永久性瘤胃瘘管的中国美利奴(新疆型)绵羊[平均体重为(32.00±1.36)kg],饲喂精粗比为30∶70的饲粮,依次采集饲喂前(0 h)和饲喂后1.5、3.0、6.0、9.0、12.0 h 6个时间点的瘤胃液,重复采集3次。分离和制备细菌、原虫破碎液,分别测定相关酶活力及谷氨酸脱氢酶体系的Km值。结果显示:1)绵羊瘤胃细菌、原虫破碎液中蛋白酶、谷丙转氨酶、谷草转氨酶和谷氨酸脱氢酶的活力随饲喂时间的延长均呈现先升高后降低的动态变化规律,总体在饲喂后1.5 h达到峰值;谷氨酸和氨含量也呈现相似的变化规律。原虫破碎液中参与蛋白质分解代谢的这4种酶的活力在各时间点均极显著高于细菌(P0.01)。2)原虫破碎液谷氨酸含量极显著高于细菌(P0.01);原虫破碎液氨含量在1.5、6.0、9.0和12.0 h显著或极显著高于细菌(P0.05或P0.01)。3)绵羊瘤胃细菌、原虫谷氨酸脱氢酶对烟酰胺腺嘌呤二核苷酸(NAD)的Km值分别为2.60×10~(-7)、1.48×10~(-7)mol/L;细菌、原虫谷氨酸脱氢酶对谷氨酸的Km值分别为8.41×10~(-6)、4.91×10~(-6)mol/L;细菌、原虫谷氨酸脱氢酶对还原型烟酰胺腺嘌呤二核苷酸(NADH)的Km值分别为3.80×10~(-8)、2.70×10~(-8)mol/L;细菌、原虫谷氨酸脱氢酶对α-酮戊二酸的Km值分别为1.16×10~(-6)、2.07×10~(-6)mol/L;细菌、原虫谷氨酸脱氢酶对氨的Km值分别为2.97×10~(-5)、1.40×10~(-5)mol/L。结果提示,总体上,绵羊瘤胃细菌、原虫中蛋白酶、谷氨酸脱氢酶、谷丙转氨酶、谷草转氨酶的活力在饲喂后1.5 h达到峰值,之后逐渐降低;绵羊瘤胃原虫中蛋白酶、谷丙转氨酶、谷草转氨酶和谷氨酸脱氢酶的活力均极显著高于细菌,原虫中蛋白质分解代谢更旺盛;瘤胃原虫中不仅存在谷氨酸转氨机制,还可能存在利用氨重新合成氨基酸的机制。 相似文献
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为探索十二指肠贾第虫(Giardia duodenalis)谷氨酸脱氢酶(glutamate dehydrogenase, GDH)在滋养体中的分布和酶动力学等生物学特性，对GDH进行了克隆并构建原核表达质粒pET-28a-GDH,然后转化至BL21(DE3) plysS感受态细胞中经IPTG诱导表达，对表达产物进行SDS-PAGE和Western blot分析，以Ni2+亲和层析纯化。对纯化的重组GDH进行体外酶动力学分析并免疫小鼠制备多克隆抗体，最后用间接免疫荧光技术对滋养体中的GDH进行定位。结果显示，贾第虫GDH的编码序列长1 350 bp,试验成功构建pET-28a-GDH重组表达质粒并诱导表达，重组蛋白约为49.7 ku,且大部分以可溶性形式存在。ELISA和Western blot结果表明，贾第虫GDH具有良好免疫原性，获得的多克隆抗体具有较强特异性。间接免疫荧光定位显示GDH主要位于滋养体的胞质中。酶动力学研究结果显示，GDH最适反应pH约为9.3、最适温度约为42℃,对L-谷氨酸和NADP+的Km值分别为4.485 mmol... 相似文献
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盐胁迫对桑树NH_4~+同化和谷氨酰胺合成酶活性的影响 总被引：2，自引：2，他引：0
以育 15 1和新一之 2个桑品种的硬枝扦插苗为材料 ,研究外源NaCl胁迫对桑树氮素代谢有关酶活性和氨积累的影响。结果表明 :NaCl胁迫显著抑制桑树的营养生长 ;可促进游离NH+4在桑树叶片中积累 ,但NH+4在根系中未见明显变化 ;受NaCl胁迫的桑树根和叶片的谷氨酰胺合成酶的活性明显下降 ,而叶片的谷氨酸脱氢酶的活性显著增加 ;桑树根系硝酸还原酶的活性受盐胁迫的刺激 ,但桑树叶片的硝酸还原酶活性明显下降 ;暗示盐分胁迫可减弱桑树经谷氨酰胺合成酶 -谷氨酸合酶循环对NH+4的同化 ,但可加强NH+4经谷氨酸脱氢酶的同化 ,并可通过影响NO-3 的还原来影响桑树的氮素营养 相似文献
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《中国兽医学报》2017,(2):327-334
为探讨日粮粗蛋白(crude protein,CP)水平对生长育肥猪肠道微生物胞内酶活性的影响,试验分别选择体质量分别为(30±2.54),(60±3.18)kg的杜×长×大三元杂交猪各18头,均采用单因子完全随机分组设计分配到CP为12%,15%,18%和10%,13%,16%的3组日粮中,每组6个重复,每个重复1头猪。预试期7d,正试期30d。在正饲期31d,每组随机挑选6头进行屠宰,采集各肠段食糜和粪便样品,研究日粮CP水平对生长育肥猪肠道微生物胞内酶活性的影响。结果表明,生长育肥猪肠道微生物酶活力均具有肠段特异性;日粮CP水平能显著影响猪肠道微生物硝酸还原酶(P<0.05)、谷氨酰胺合成酶(P<0.05)、谷草转氨酶(P<0.05)、谷丙转氨酶(P<0.05)、乳酸脱氢酶(P<0.05)、尿素酶(P<0.05)及蛋白酶(P<0.05)水平;日粮CP水平对肠道微生物的蛋氨酸、谷氨酸、精氨酸、酪氨酸、色氨酸脱羧酶活力影响显著(P<0.05);日粮CP水平显著影响肠道微生物酶的活力,是肠道微生物对氨基酸差异性降解和动物对蛋白质利用率低的原因之一。 相似文献
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反刍动物能很大程度上依靠瘤胃微生物来消化利用饲料中的植酸盐,但目前国内有关瘤胃微生物植酸酶的研究报道较少.本文对瘤胃微生物植酸酶的主要来源和活性进行了综述分析,发现主要有5种瘤胃细菌能分泌植酸酶,其中,反刍兽新月形单胞菌(Selenomonas ruminantium)分泌的植酸酶活性最高(199.1～703.6 U·mL-1)；根据酶促反应的最适pH和催化机理,可将瘤胃微生物植酸酶分别归类为酸性植酸酶和半胱氨酸磷酸酶植酸酶；根据植酸酶水解植酸时的立体专一性,发现瘤胃微生物植酸酶中可能同时存在3-植酸酶和5-植酸酶2类植酸酶.最后本文论述了瘤胃微生物植酸酶酶促反应动力学特性,发现反刍兽新月形单胞菌植酸酶的最适pH为4.0～5.5、最适温度为50～55℃、对植酸和ATP表现出较强的活性；埃氏巨形球菌植酸酶的最适pH为5,0、最适温度为60℃、仅对植酸表现出活性. 相似文献
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《中国兽医学报》2019,(5):982-989
旨在研究断奶仔猪肠道微生物氮代谢相关酶活对日粮粗蛋白(crude protein,CP)水平响应规律。选择54头体质量为(5.55±0.49) kg的28日龄杜洛克×长白×大约克断奶仔猪,采用单因子完全随机设计分配到CP为20%、17%、14%的3组日粮中,每组18个重复,每个重复1头猪。预试期7 d,正试期45 d。分别在正试期10,25及45 d,每组随机挑选6头进行屠宰,采集各肠段食糜和粪便样品分离微生物,测定肠道微生物氮代谢相关酶活力。结果表明,仔猪肠道微生物氮代谢相关酶酶活力具有肠段特异性,日粮高CP水平能显著提高肠道微生物硝酸还原酶(P0.05)、腺苷脱氨酶(P0.05)、谷氨酰胺合成酶(P0.05)、尿素酶(P0.05)、谷丙转氨酶(P0.05)及蛋白酶(P0.05)。肠道微生物的蛋氨酸、组氨酸、赖氨酸、精氨酸、甘氨酸、酪氨酸、色氨酸、谷氨酸脱羧酶活力对日粮高CP水平响应明显(P0.05)。消化道微生物氮代谢相关酶活力对蛋白质水平的响应规律性解析了当前猪蛋白质利用率低的原因。 相似文献
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高温胁迫下不同氮肥处理对高羊茅氮代谢的影响 总被引：3，自引：1，他引：2
采用盆栽试验,研究了不同氮肥（NO3--N、NH4+-N、NH4NO3-N）处理对高羊茅品种凌志Festuca arundinacea cv. Barlexas在高温38/30 ℃（昼/夜）胁迫下叶片内硝态氮含量、铵态氮含量以及参与氮同化和代谢过程的主要酶活性的影响。结果表明：随着胁迫时间的延长,经过不同氮肥处理的植株中硝态氮含量先升后降,而铵态氮含量呈上升趋势,其中NH4NO3-N处理株的硝态氮含量最高,铵态氮含量最低,NH4+-N处理株的铵态氮含量最高;不同处理株中的硝酸还原酶（NR）活性呈下降趋势,而谷氨酰胺合成酶(GS)和谷氨酸合成酶（GOGAT）活性均呈先升后降的趋势,总体上,NH4NO3-N处理株的NR、GS和GOGAT活性最高;NH4NO3-N和NO3--N处理株的谷氨酸脱氢酶（GDH）活性均呈下降趋势,而NH4+-N处理株的GDH活性呈上升趋势。在试验条件下,经NH4NO3-N处理的高羊茅受氧化胁迫程度最小,耐热性最好,有利于高羊茅越夏和延长其绿期。 相似文献
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K Holovská V Lenártová J Koppel I Havassy P Javorsky E Rybosová 《Veterinární medicína》1991,36(10):625-632
Developmental dynamics was investigated in the activity of glutamate dehydrogenase (GDH, E.C. 1.4.1.2.-4) and glutamine synthetase (GS, E.C. 6.3.1.2) in different parts of the digestive tract of lambs, in dependence on the age from 10 to 90 days; the goal of these investigations was to elucidate in greater detail the role of the above enzymes in nitrogen metabolism. The activity of GDH, and of the coenzymes NADH and NADPH, was followed in the digesta because simple organisms (bacteria, fungi, plants) have two glutamate dehydrogenases: they differ from each other by coenzyme specificity, unlike GDH from animal sources which can utilize both NADH coenzyme and NADPH coenzyme (Fahien et al., 1965; Frieden, 1964). The following activities of GDH and GS were found out in trials with lambs at the age of 10, 20, 30, 40 and 90 days, as to the different parts of digestive tract: in the tissues of rumen, omasum, reticulum, spleen, duodenum, jejunum, ileum, int. caecum and colon the activity of GDH (NADH) varied from 0.031 to 0.305 nkat/mg dry matter, in the digesta from 0 to 2.92 nkat/mg dry matter. An investigation of GDH (NADH, NADPH) dynamics in the digesta of lambs showed the relatively high activity of GDH (NADH) in the digesta of colon at the age of 10 days and that of GDH (NADPH) in the digesta of int. caecum. The activity of GDH (NADH) was also found to be high in the digesta of int. caecum at the age of 20 days. In that period the activity of GDH (NADH, NADPH) in the digesta of rumen, omasum and reticulum was zero.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
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The use of selected plasma enzyme activities for the diagnosis of fatty liver-hemorrhagic syndrome in laying hens 总被引：8，自引：0，他引：8
Profiles of plasma enzymes were compared in two strains of single comb white leghorn laying hens, a normal commercial strain and strain UCD-003, which is highly susceptible to fatty liver-hemorrhagic syndrome. Plasma activity of lactate dehydrogenase (LDH), glutamate dehydrogenase (GDH), aspartate aminotransferase (AST), alanine aminotransferase (ALT), and creatine kinase (CK) averaged 194 +/- 27, 4.0 +/- 2.8, 146 +/- 20, 1.0 +/- 1.0, and 1041 +/- 268 U/liter, respectively in normal birds. Activities of LDH, GDH, AST, and ALT, but not CK, were significantly higher in UCD-003 than in normal hens. A bimodal distribution of activities of all enzymes was found in the UCD-003 hens, with some birds showing activities comparable with those of the normal hens and others with values that were 2-10 times greater than those found in normal hens. These results are consistent with the extensive hepatic lesions observed in the UCD-003 strain of birds. Average gross hemorrhagic scores from visual inspection (scale of 0-3) were 0.28 +/- 0.45 in normal birds and 1.63 +/- 0.94 in the UCD-003 birds. Even though no clear relationship was found between plasma enzyme activities and the extent of liver hemorrhage in individual birds, the UCD-003 hens consistently had average values significantly higher for plasma enzymes that indicate liver damage. The results suggest that measurement of enzyme activities indicative of liver damage in birds, particularly AST, LDH, and GDH, is a valuable tool in the diagnosis of fatty liver-hemorrhagic syndrome in a flock of layers. 相似文献
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The clearance of glutamate dehydrogenase from plasma was measured weekly for three weeks in three dry and three lactating cows. The clearance was exponential with a mean clearance constant of 0.0488/j and means (+/- SE) half-life of 14.2 (+/- 0.77) h. There were variations among cows and among measurements in the same cow but there was no difference between the mean half-lives of GDH in lactating and dry cows. Because of the long half-life the small variations among individual cows are unlikely to affect the interpretation of increased plasma GDH activity. 相似文献
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为揭示NO对渗透胁迫下紫花苜蓿含氮化合物及氮代谢过程的调控机制,以紫花苜蓿为材料,通过外源施加NO供体硝普钠（SNP）和NO清除剂（c-PTIO）对紫花苜蓿种子及幼苗进行处理,研究NO在PEG模拟的渗透胁迫下紫花苜蓿氮类化合物含量及氮代谢关键酶活性变化规律中的影响。结果表明：与PEG处理相比,外施0.1 mmol·L-1 SNP,紫花苜蓿萌发期及幼苗期可溶性蛋白含量分别平均提高了 16.91%和8.87%,游离氨基酸含量平均降低14.92%和0.68%;硝酸还原酶（NR）、谷氨酰胺合成酶（GS）及谷氨酸合成酶（GOGAT）活性在萌发期分别提高了23.85%,47.87%,60.06%,在幼苗期分别提高了16.31%,23.56%,72.89%;而萌发期和幼苗期谷氨酸脱氢酶（GDH）活性分别降低了35.89%和33.35%。PEG胁迫下添加c-PTIO加剧了PEG胁迫对紫花苜蓿含氮物质积累的抑制,对NR,GS,GOGAT活性的影响差异不显著,GDH活性在萌发期及幼苗期分别平均升高了18.26%和4.62%。由此说明,外源NO通过调节氮代谢关键酶活性促进PEG胁迫下紫花苜蓿氮素向蛋白质合成的方向转运积累,提高紫花苜蓿抗旱性。 相似文献
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In vitro alterations induced by a 10 micrograms/ml and 50 micrograms/ml dose each of thiophenate and fenbendazole on the absorptive surfaces of Haemonchus contortus (Nematoda: Trichostrongylidae) were studied. The most significant changes were induced in the gut epithelium. Alkaline phosphatase and adenosine triphosphatase activities were decreased, succinic dehydrogenase activity was increased, while acid phosphatase and glucose-6-phosphatase were completely lost from the intestinal epithelium after treatment with either of the drugs. A stimulatory effect of these two anthelmintics was observe on lactic dehydrogenase and reduced nicotinamide adenine dinucleotide diaphorase distribution. Thiophenate caused an increase in the activities of glutamate dehydrogenase (GDH), glucose-6-phosphate dehydrogenase (G-6-PD) and nonspecific esterases and a decrease in reduced nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-D) activity. Fenbendazole treatment led to the inhibition of GDH, while G-6-PD, NADPH-D, cytochrome oxidase, monoamine oxidase and nonspecific esterase activity remained unaltered in the epithelium. 相似文献
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Tasuke Ando Yoko Sugawara Ryohei Nishio Miho Murakami Emiko Isogai Hiroshi Yoneyama 《Animal Science Journal》2017,88(7):1027-1033
Streptococcus bovis, an etiologic agent of rumen acidosis in cattle, is a rumen bacterium that can grow in a chemically defined medium containing ammonia as a sole source of nitrogen. To understand its ability to assimilate inorganic ammonia, we focused on the function of glutamate dehydrogenase. In order to identify the gene encoding this enzyme, we first amplified an internal region of the gene by using degenerate primers corresponding to hexameric family I and NAD(P)+ binding motifs. Subsequently, inverse PCR was used to identify the whole gene, comprising an open reading frame of 1350 bp that encodes 449 amino acid residues that appear to have the substrate binding site of glutamate dehydrogenase observed in other organisms. Upon introduction of a recombinant plasmid harboring the gene into an Escherichia coli glutamate auxotroph lacking glutamate dehydrogenase and glutamate synthase, the transformants gained the ability to grow on minimal medium without glutamate supplementation. When cell extracts of the transformant were resolved by blue native polyacrylamide gel electrophoresis followed by activity staining, a single protein band appeared that corresponded to the size of S. bovis glutamate dehydrogenase. Based on these results, we concluded that the gene obtained encodes glutamate dehydrogenase in S. bovis. 相似文献
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对猪链球菌2型、7型、9型的标准菌株和7株猪链球菌2型分离菌的谷氨酸脱氢酶(GDH)基因进行PCR扩增,经回收纯化后克隆到PMD18-T载体,筛选阳性克隆菌后测序并对其进行序列分析。结果,PCR扩增出1300bp左右的片段,包括GDH基因的整个开放阅读框,而测序结果表明,其序列与GenBank中的猪链球菌GDH基因序列一致,进一步序列分析表明,GDH的核苷酸序列在相同血清型之间同源性高达99%以上,而在不同血清型之间的同源型也达到了96%以上,而其氨基酸序列同源性则都在99%以上,且都具有GDH1型家族的功能区域。说明猪链球菌GDH基因及其蛋白具有高度的保守性,为进一步研究与应用提供了重要依据。 相似文献