Robert A. Weisberg

Bio: Robert A. Weisberg is an academic researcher from National Institutes of Health. The author has contributed to research in topics: Bacteriophage & RNA polymerase. The author has an hindex of 36, co-authored 76 publications receiving 4135 citations. Previous affiliations of Robert A. Weisberg include Institute of Cancer Research & Tel Aviv University.

Overproduction of Escherichia coli integration host factor, a protein with nonidentical subunits.

Abstract: Integration host factor (IHF) is a small, basic protein that is needed for efficient recombination of bacteriophage lambda, as well as for other host and viral functions. We have constructed strains in which the two subunits of IHF, encoded by the himA and hip genes of Escherichia coli, are expressed under the control of the lambda rho L promoter. Separate overexpression of himA and hip led to the production of unstable and insoluble peptides, respectively. In contrast, the overexpression of both genes conjointly led to the accumulation of large amounts of active IHF. Extracts of such cells provided the starting material for a rapid purification procedure that results in milligram quantities of apparently homogeneous IHF.

Site-specific Recombination in Phage Lambda

Abstract: INTRODUCTION Cells lysogenic for λ carry a quiescent form of the viral chromosome called prophage The prophage differs from the DNA of viral particles in two important ways: (1) The ends of the prophage and of the viral particle DNA are at different points in the nucleotide sequence, and (2) the prophage ends are covalently joined to host DNA Campbell (1962) proposed that viral particle DNA is converted to prophage by the joining of the ends followed by insertion of the resulting circular molecule into the host DNA Insertion occurs by reciprocal recombination at specific sites (attachment or att sites) in each chromosome (Fig 1) This proposal has received extensive experimental support and, indeed, was generally accepted when the first edition of this book was written (see Gottesman and Weisberg 1971) A stable lysogen is formed when an infecting viral particle succeeds both in repressing lytic functions and in inserting its DNA The inserted prophage is then replicated as part of the bacterial chromosome In the rare event that repression is not followed by insertion (abortive lysogeny), the viral chromosome cannot replicate and so is lost by dilution as the host cell divides Excision of the prophage from the chromosome is rare during normal bacterial growth but occurs readily following repressor inactivation If the repressor is inactivated only briefly (abortive or transient derepression), prophage excision can occur without lytic growth The excised DNA is then frequently lost as the cell divides (prophage curing) Intricate controls ensure that insertion occurs only

Steroid Receptor Regulated Transcription of Specific Genes and Gene Networks

Abstract: PERSPECTIVES AND SUMMARY . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 210 ORIGINS OF CONSERVED GENE REGULATORS . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . 211 Steroids . . . . .. . . . . . . . . . . . . . . . . . . . .. . . .. . . . . . . . . . . . .. . . . . .. . . . . . . . . . . . . . .... . . . . . . .. . . . . . . . .. . . . . . . .. . 211 Steroid Receptors .... " . . . . . . . . . . . . . . . . . . . . . " . . . . . . . . . . . . . . . . . . . . . . . . "" . . . . . . . . . . . . . . . . . . . . . . " 211 FUNCTIONAL ASPECTS OF RECEPTOR STRUCTURE" . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 213 Activation and Intracellular Localization ... . ". . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 213 Steroid Binding .. ...... , . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . " . . . . . . . . . . . . . . . . . . , 214 Nonspecific DNA Binding . .. . ..... ....... . ....... " . . . . . . . . .. . . . . . . . . . . . . . . . . " 215 Functional Heterogeneity 216 SELECTIVE RECEPTOR:DNA INTERACTIONS ... . . .... .. .... . .... . 21 7 Receptor Binding at Genomic Sites of Action . " . . . . . . . " 218 Detection of Specific DNA Binding Sites """""",,, , , """""'" . . . . . . . . . . . . . . . . . . . . . . . 21 8 DNA Sequences Bound by Steroid Receptors .. . ......... . ..... . ..... . ..... . ... , 221 LOCALIZATION OF STEROID RESPONSE ELEMENTS (REs) . .... " . . . . . . . . . 222 Deletion Mapping .. . ..... ......... . ....... . " . . . . . . . . . . . . . . . . . . . . . ""." . . .... . . . 223 RE-Promoter Fusions .... ......... . " . . . . . . . . . . . . . . . . . ". . . . . . . . . . . . . . . . . . 223 In Vitro Mutagenesis" ...... . ........ ......... .. " . . . . . . . . . . . . . . . . . "" . . . . . . " . . . . . . . . . . . . . . . . " 224 TRANSCRIPTIONAL ENHANCEMENT BY RECEPTOR:RE COMPLEXES " " "'" ' ' 224 Glucocorticoid-Dependent Enhancement 225 Detection and Characterization of Enhancers. . .. . . . . . . .... . . .... . . . . . . .. . . . . . . .... . . ... . . . . 225 EFFECTS OF RECEPTOR BINDING ON GENOME STRUCTURE ... " . . ". . . . . . . . . . . . . . 227 Cytological and Nuclease Probes .. . . """""" . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . " . . . . . . . . . 227 Chromosomal Protein Alterations 229 DNA-Structure Alterations 230 New Experimental Approaches . . . . . ... """ . . ,, .. " . . . . ,,". . . . . . . . . . . . . . . . . . . . . . . . . 232 HIGHER-ORDER CONTROL . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . .. 233 Gene Networks ..... ...... ........ ....... . " . . . . . . . . , . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ". . . . . . . . . 234 Multifactor Regulation ...... ... ,. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 235

An efficient recombination system for chromosome engineering in Escherichia coli

Abstract: A recombination system has been developed for efficient chromosome engineering in Escherichia coli by using electroporated linear DNA. A defective lambda prophage supplies functions that protect and recombine an electroporated linear DNA substrate in the bacterial cell. The use of recombination eliminates the requirement for standard cloning as all novel joints are engineered by chemical synthesis in vitro and the linear DNA is efficiently recombined into place in vivo. The technology and manipulations required are simple and straightforward. A temperature-dependent repressor tightly controls prophage expression, and, thus, recombination functions can be transiently supplied by shifting cultures to 42 degrees C for 15 min. The efficient prophage recombination system does not require host RecA function and depends primarily on Exo, Beta, and Gam functions expressed from the defective lambda prophage. The defective prophage can be moved to other strains and can be easily removed from any strain. Gene disruptions and modifications of both the bacterial chromosome and bacterial plasmids are possible. This system will be especially useful for the engineering of large bacterial plasmids such as those from bacterial artificial chromosome libraries.