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91.
Revertants and Secondary arom-2 Mutants Induced in Non-Complementing Mutants in the arom Gene Cluster of Neurospora Crassa 总被引:7,自引:0,他引:7 下载免费PDF全文
Extensive genetical and biochemical studies have been performed with revertants and secondary arom-2 mutants induced in two different primary non-complementing mutants which map within the arom gene cluster of Neurospora crassa. These studies indicate that mutant M54 but not M25 can revert by super-suppressor mutations in unlinked genes, thus confirming previous evidence that M54 contains a nonsense codon. At least three new super suppressors of M54 have been detected. All four super suppressors (including one previously detected) when combined with M54 result in high levels of all five of the arom enzymic activities in the form of arom multienzyme complexes very similar to (but not necessarily identical with) that in wild type (WT).-Evidence has also been obtained that the two non-complementing mutants can yield revertants which appear to result from true back mutations and produce arom aggregates essentially indistinguishable from that of WT. In addition, M25, but not M54, when plated on quinic acid yields revertants (secondary mutants) some of which are phenotypically indistinguishable from arom-2 primary mutants and others of which, although also mapping within the arom-2 gene, exhibit unusual properties. Genetic evidence indicates that the M25 secondary mutants are localized within the arom-2 gene, but that they arise from mutational events more complex than ones resulting in single base pair changes in the M25 codon.-The recovery of secondary arom-2 mutants as revertants of non-complementing arom mutants provides strong evidence, independent of earlier recombination data, that non-complementing arom mutants are located within the arom-2 structural gene of the arom gene cluster. In addition, the occurrence and characteristics of these secondary arom-2 mutants provide strong evidence, independent of the results with nonsense suppressors, that the arom gene cluster is transcribed, beginning with the arom-2 gene, as a single polycistronic messenger ribonucleic acid (mRNA) molecule which is subsequently translated into the arom multienzyme complex. 相似文献
92.
The coexistence and coevolution of sexual and asexual species under resource competition are explored with three models: a nongenetic ecological model, a model including single locus genetics, and a quantitative-genetic model. The basic assumption underlying all three models is that genetic differences are translated into ecological differences. Hence if sexual species are genetically more variable, they will be ecologically more variable. Under classical competition theory, this increased ecological variability can, in many cases, be an advantage to individual sexual genotypes and to the sexual species as a whole. The purpose of this paper is to determine the conditions when this advantage will outway three disadvantages of sexuality: the costs of males, of segregation, and of the additive component of recombination. All three models reach similar conclusions. Although asexuality confers an advantage, it is much less than a two-fold advantage because minor increases in the overall species niche width of the sexual species will offset the reproductive advantage of the asexual species. This occurs for two reasons. First, an increase in species niche width increases the resource base of the sexual species. Second, to the extent that the increase in niche width is due to increased differences between individuals, a reduction in intraspecific competition will result. This is not to imply that the sexual species will always win. The prime conditions that enable sexual species to stably coexist with or even supplant an asexual sister species are:
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A 4'-oxidized abasic site (X) has been synthesized in a defined duplex DNA sequence, 5'-d(CCAAAGXACCGGG)-3'/3'-d(GGTTTCATGGCCC)-5' (1). Its structure has been determined by two-dimensional NMR methods, molecular modeling, and molecular dynamics simulations. 1 is globally B-form with the base (A) opposite X intrahelical and well-stacked. Only the alpha anomer of X is observed, and the abasic site deoxyribose is largely intrahelical. These results are compared with a normal abasic site (Y) in the same sequence context (2). Y is composed of a 60:40 mixture of alpha and beta anomers (2alpha and 2beta). In both 2alpha and 2beta, the base (A) opposite Y is intrahelical and well-stacked and the abasic site deoxyribose is predominantly extrahelical, consistent with the reported structures of the normal abasic site in a similar sequence context [Hoehn, S. T., Turner, C. J., and Stubbe, J. (2001) Nucleic Acids Res. 29, 3413-3423]. Molecular dynamics simulations reveal that the normal abasic site appears to be conformationally more flexible than the 4'-oxidized abasic site. The importance of the structure and flexibility of the abasic site in the recognition by the DNA repair enzyme Ape1 is discussed. 相似文献
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Morton MF Liu PQ Reik A de la Rosa R Mendel M Li XY Case C Pabo C Moreno V Pyati J Shankley NP 《Regulatory peptides》2005,129(1-3):227-232
Designed zinc finger proteins (ZFPs) regulate expression of target genes when coupled to activator or repressor domains. Transfection of ZFPs into cell lines can create expression systems where the targeted endogenous gene is transcribed and the protein of interest can be investigated in its own cellular context. Here we describe the pharmacological investigation of an expression system generated using CCK2 receptor-selective ZFPs transfected into human embryonic kidney cells (HEKZFP system). The receptors expressed in this system, in response to ZFP expression, were functional in calcium mobilization studies and the potency of the agonists investigated was consistent with their action at CCK2 receptors (CCK-8S pA50 = 9.05+/-0.11, pentagastrin pA50 = 9.11+/-0.13). In addition, binding studies were conducted using [125I]-BH-CCK-8S as radioligand. The saturation binding analysis of this radioligand was consistent with a single population of high affinity CCK receptors (pK(D) = 10.24). Competition studies were also conducted using a number of previously well-characterized CCK-receptor selective ligands; JB93182, YF476, PD-134,308, SR27897, dexloxiglumide, L-365,260 and L-364,718. Overall, the estimated affinity values for these ligands were consistent with their interaction at CCK2 receptors. Therefore, CCK2 receptors up-regulated using zinc finger protein technology can provide an alternative to standard transfection techniques for the pharmacological analysis of compounds. 相似文献
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Like temperature and pressure, the solution pH is an important thermodynamic variable that is commonly varied in experiments and is used by cells to influence biochemical function. It is now becoming feasible to carry out practical molecular dynamics simulations that mimic the thermodynamics of such experiments, by allowing proton transfer between the system of interest and a hypothetical bath of protons at a given pH. These are demanding calculations, because the energetics of charge changes upon protonation or deprotonation must be accurately modeled, and because such simulations must sample both molecular configurations and the large number of protonation states that are possible for a molecule with many titrating sites. 相似文献
100.