GENETIC AND BIOCHEMICAL ANALYSIS OF AD 5 REGION 2

目的是了解控制机制 在真核基因表达的调节中起作用，使用腺病毒 5型（Ad 5）早期区域2A作为模型系统。 的基因产物 早期区域2A是72，000道尔顿的单链DNA结合蛋白 (DBP)参与调节病毒基因表达的基因， 方法：（1）DNA合成的起始和延伸步骤，（2） 调节几个Ad 5早期基因的转录，（3） 早期mRNA的稳定性;（4）病毒的宿主范围。 当前的目标是在DBP域中精确定义 参与蛋白质的每一种功能。 为了做到这一点，我们 将通过位点特异性诱变技术在DBP中产生突变 并分析突变的DBP与单链DNA结合的能力 DNA，并帮助腺相关病毒在人体细胞中复制。 深入探讨蛋白质的功能特性将是 在将突变基因重新引入病毒后进行。

The objective is to gain an understanding of the control mechanisms operative in the regulation of eukaryotic gene expression, using adenovirus type 5 (Ad5) early region 2A as the model system. The gene product of early region 2A is a 72,000 dalton single-stranded DNA-binding protein (DBP) that is involved in regulating viral gene expression in at least four ways: (1) the initiation and elongation steps of DNA synthesis, (2) the regulation of transcription of several of the Ad5 early genes, (3) stability of early mRNAs, and (4) the host-range of the virus. The immediate goal is to precisely define within the DBP domains that are involved in each of the functions of the protein. To accomplish this, we will generate mutations in the DBP by site-specific mutagenesis techniques and analyze the mutant DBPs for their ability to bind to single-stranded DNA, and to help adeno-associated virus replicate in human cells. In-depth probing of the functional properties of the protein will be performed following re-introduction of the mutated gene back into the virus.