【作者】 张晓红；

【导师】 刘丽；

【作者基本信息】 广州医学院 ， 儿科学， 2010， 硕士

【摘要】 背景酮体是能量从肝脏向肝外组织转移的主要载体[1],主要在肝脏生成。体内酮体浓度增加时即发生酮症酸中毒。琥珀酰辅酶A转硫酶(SCOT)是酮体能量代谢的关键酶,位于多个组织的线粒体内,其作用是将琥珀酰辅酶A的辅酶A基团转移至乙酰乙酸从而催化乙酰乙酰辅酶A的形成,这一步骤是酮体利用的第一步,随后乙酰乙酰辅酶A进一步断裂成两分子的乙酰辅酶A进入三羧酸循环。研究表明:SCOT催化乙酰乙酰辅酶A生成的具体机制是与辅酶A的巯基生成硫酯并以非共价键的形式连接辅酶A的ADP基团。硫酯形成的部位位于SCOT肽链第344位谷氨酸的羧基上。SCOT缺乏导致发作性的酮症酸中毒,为常染色体隐形遗传,大多数情况下作为儿童酮症酸中毒鉴别诊断的一部分。与大多数有机酸血症不同的是,除乙酰乙酸和3-羟基丁酸等酮体增加外,这类病人的血和(或)尿液中无其他诊断性的代谢产物出现。自1972年J.T.Tildon等报道的第一例SCOT缺陷病人至今,只发现18例先证者[1-12]]和10种突变类型[3,5,6,8-12]。目的探讨解酮障碍SCOT基因突变类型以及解酮障碍患者基因型和临床表型之间的关系。方法提取3例临床高度SCOT基因缺陷所致解酮障碍患儿以及50例正常对照儿童外周血总RNA,进行RT-PCR反应,扩增整个SCOT cDNA编码区片段并克隆测序,对RNA检测异常的患儿进行DNA检测;观察基因突变患儿的临床表现。结果3例患者中,1例发现12外显子的跳跃现象,DNA水平检测发现12外显子第48位碱基发生替代(c1147G>A)。mRNA前体二级结构分析显示12外显子的这种替代确实改变了正常序列的mRNA前体二级结构,ESE分析提示碱基替代位点所在的这段嘌呤富集区作为ESE的可能性并不大。其余2例未发现突变。SCOT基因12外显子跳跃的患儿表现为感染诱发的反复发作的严重酮症酸中毒,经积极抢救症状能缓解,发作间歇期患儿同正常儿童,生长发育正常。结论SCOT基因12外显子48位碱基的替代是否通过改变mRNA前体的二级结构而导致12外显子跳跃仍有待进一步的证实。该突变在国内外未见报道。对SCOT基因12外显子跳跃的患儿应避免感染,发作时应进行积极救治。更多还原

【Abstract】 Background Ketone body are major vectors of energy transfer from liver to extrahepatic tissues[1] which are mainly produced in liver. Ketoscidosis occurs commonly under the condition of a marked increase in ketone body concentration. Succinyl CoA: 3-oxoacid CoA transferase(SCOT) is the key enzyme of ketone body utilization, existing in the mitochondrial matrix of multiple organs. SCOT catalyzes the reversible transfer of a CoA moiety from succinyl CoA to the ketone body acetoacetate, the first step of ketone- body utilization. Then, acetoacetyl CoA cleavage to two acetyl CoA molecules which are capable of entering the tricarboxylic acid cycle. Elegant studies revealed that, the special mechanism of SCOT catalyzing the formation of acetoacetyl CoA is generating the thioester with coenzyme A and binding the ADP groups of coenzyme A in the form of non–covalent bond. The formation of thioester occurs in on the glutamate residue344 of SCOT peptide chain. Hereditary deficiency of SCOT is an autosomal recessive inborn error and is part of the differential diagnosis of childhood ketoacidosis , a frequently occurring condition. In contrast with most organic acidemias, no diagnostic metabolites are observed in blood and urine samples from SCOT-deficient patients although the ketone bodies acetoacetate and 3-hydroxybutyrate are elevated. Since the first description of SCOT deficiency, only 18 [1-12]]affedted probands and 10 mutations have been reported[3,5,6,8-12].Objective To explore the mutations types of Chinese SCOT-deficient patient and the relationship between the genotype and phenotype.Method Total RNA was extracted from peripheral blood of clinical highly suspected children and 50 health children as control, the whole coding sequence of SCOT cDNA was amplified by RT-PCR followed by T-Clone and sequencing.DNA was teseted if RNA was tested abnormal; observed the clinical manifestions of the child who had SCOT gene mutations.Result One of there patients was found with skipping of exon 12, detection in DNA level showed a single-base substitution(c1147G>A)in exon 12. The pre-mRNA secondary structure analysis revealed a change of pre-mRNA secondary structure after substitution while the ESE analysis told us that the purine-rich region was unlikely acting as ESEs. The other two were found no mutations. The child who had 12 exon skip of SCOT gene mainly manifested as recurrent infection induced severe ketoacidosis, but the symptoms could released by active salvage, and the patient could be as normal as other children during the intermittent episodes.Conclusion Whether the substitution of exon 12(c1147G>A)resulted in the skipping of 12 by disrupting the pre-RNA secondary structure needs to be further confirmed. Thjs mutation had not been reported before. Infection should be avoided if the child has this kind of mution, and treatment should be active.更多还原