Spontaneous inactivation of enkephalin.

The role of isoleucyl-, valyl-, and leucyl-tRNA synthetases in attenuation of the $\text{ilvEDA}$ operon was examined. The results indicate that the activities of isoleucyl- and valyl-tRNA synthetases are necessary to maintain attenuation of the $\text{ilvEDA}$ operon. Leucyl-tRNA synthetase activity is nonessential for attenuation. These studies imply that uncharged tRNA^Ile and tRNA^Val each may cause deattenuation.     

Localization of the lactose permease protein(s) in the E. coli envelope.

The amino acid double labeling technique was used to identify and localize membrane-bound lactose operon proteins in $\text{E.}\text{coli}$. Both the “M” protein, thought to be the $\text{y}$ gene product, and a polypeptide of MW ～15,000 appeared in the membrane following $\text{lac}$ operon induction. The amounts of these two proteins were approximately equal.The inner and outer membrane layers of the cell envelope were separated by sucrose density gradient centrifugation or by selective solubilization of inner membranes with the detergent Sarkosyl. When gentle lysis conditions were employed to prepare membrane vesicles, both $\text{lac}$ induced proteins fractionated with the inner membrane. However, the “M” protein was more easily randomized in the envelope structure by sonication than the 15,000 dalton component or an inner membrane marker enzyme.     

Regulation of the Escherichiacoli tryptophan operon by readthrough of UGA termination codons

The regulation of the synthesis of $\text{trp}$ operon enzymes was studied in streptomycin-resistant $\text{Escherichia}\text{coli}$ mutants temperature-sensitive for UGA suppression by normal tRNA^Trp. Our mutants carry a $\text{trp}\text{R}{}^{\mathrm{+}}$ allele that when transferred to a different genetic background causes repression of trp operon enzyme synthesis at both low (35°C) and high (42°C) temperatures; however, in our mutants with an excess of tryptophan and at increased temperatures $\text{trp}$ enzyme synthesis is derepressed. Based on our results and the sequence data of the $\text{trp}$R gene [Singleton et al. (1980) Nucleic Acids Res., 8, 1551–1560], we offer a model for the involvement of the limited misreading of UGA codons by normal charged tRNA^Trp in the autogenous regulation of the $\text{trp}$R gene expression. The UGA readthrough process may be a regulatory amplifier of the effect of tryptophan starvation.     

In vitro formation of assimilatory nitrate reductase: presence of the constitutive component in bacteria

Cell-free extracts of a selected group of bacteria which are capable of metabolyzing dinitrogen and/or nitrate contain a soluble form of the constitutive component which is active in the $\text{in}\text{vitro}$ formation of NADPH-nitrate reductase when mixed with extracts of $\text{N.}\text{crassa}\text{,}\text{nit}\text{-}\text{1}$. The constitutive component in these extracts is dialyzable and is insensitive to trypsin and protease. The constitutive component which substitutes for the absence of the $\text{nit}\text{-}\text{1}\text{gene product in the}\text{in}\text{vitro}$ formation of NADPH-nitrate reductase is postulated to be a low molecular weight cofactor or polypeptide and is shown to be present in a number of unrelated bacteria.     

Use of lac operon fusions to isolate Escherichia coli mutants with altered expression of the fumarate reductase system in response to substrate and respiratory controls.

Strains of $\text{E}\text{.}\text{coli}$ with fusions between the $\text{lac}$ structural genes and the promoter region of the fumarate reductase system were constructed from a parental strain deleted in the native $\text{lac}$ operon. Like fumarate reductase in wild-type cells, β-galactosidase in these fusion strains is inducible by fumarate, but only under anaerobic conditions. From one of these strains, three classes of mutants altered in the expression of the hybrid operon were isolated. By anaerobic selection for growth on lactose in the absence of fumarate, mutants that synthesize β-galactosidase constitutively both aerobically and anaerobically were obtained. By aerobic selection for growth on lactose in the presence of fumarate, mutants that are inducible in the enzyme both aerobically and anaerobically and mutants that are inducible in the enzyme only aerobically were obtained. The regulatory behaviors of the mutants studied suggest that substrate and respiratory control of the expression of the fumarate reductase complex are mechanistically connected.     

The separation of phage promoter from bacterial lac promoter for beta-galactosidase expression in transducing phage lambda plac5.

The replication defective transducing phage λp$\text{lac}\text{5}\text{O}\text{29}\text{P}$3 carries a portion of the $\text{E.}\text{coli}\text{lac}$ operon in the $\text{b}$2 region of the lambda phage. This $\text{lac}$ operon segment contains the $\text{lac}$ promoter, the $\text{lac}$ operator, and the β-galactosidase $\text{z}$ gene, but does not contain the $\text{lac}$ repressor $\text{i}$ gene. The $\text{z}$ gene can be expressed from both the inserted $\text{lac}$ promoter and the phage promoter. When $\text{E.}\text{coli}$ strain 594 ($\text{z}$^?, $\text{i}$⁺) or JC6256 (Δ$\text{lac}$) is infected by λp$\text{lac}\text{5}\text{O}\text{29}\text{P}$3 in the absence of additional cyclic AMP, β-galactosidase synthesis is shown to be expressed from the phage promoter. When 594 (λ⁺) or JC6256 (λ⁺) is infected by λp$\text{lac}\text{5}\text{O}\text{29}\text{P}$3 in the presence of additional cyclic AMP and IPTG, β-galactosidase synthesis is shown to be expressed from the inserted $\text{lac}$ promoter.The ability to separate the phage promoter from the inserted $\text{lac}$ promoter for β-galactosidase expression will simplify the interpretation whenever λp$\text{lac}$5 is used.     

Identification of an Escherichia coli inner membrane polypeptide specified by a lambda-tonB transducing.

Analysis by polyacrylamide gel electrophoresis of the proteins coded by a λ$\text{ton}$B transducing phage, after infection of UV-irradiated bacteria, revealed the presence of at least 7 new polypeptides. Three of these were identified as proteins of the $\text{trp}$ operon whilst three others were deleted by a spontaneous mutation in the $\text{ton}$B region carried by the phage. A single polypeptide, molecular weight 40,000 was absent from a phage carrying a proflavine induced mutation in $\text{ton}$B. We conclude that this protein, which was localised in the inner membrane by sarkosyl fractionation of the envelope, is the $\text{tonB}$ product.     

The ilvEDA operon of Escherichia coli K12 encodes only one valine-alpha-ketoglutarate transaminase activity.

Transaminase B of $\text{E. coli}$ K12 was purified to apparent homogeneity as measured by SDS acrylamide gel electrophoresis, immunoelectrophoresis, and amino terminal sequence analysis. The valine- and isoleucine-α-ketoglutarate dependent transaminase activities of pure enzyme as well as crude extracts were characterized by immunologic and kinetic methods. The data disprove the existence of a separate valine-α-ketoglutarate transaminase within the $\text{ilvEDA}$ operon.     

Localization of translation initiation in the message for E. coli UDP-galactose 4-epimerase: the amino terminal sequence

The amino acid sequence of $\text{E.}$$\text{coli}$ UDP-galactose 4-epimerase has been determined through the amino-terminal 28-amino acid residues using an automated protein sequenator. Alignment of UDP-galactose operon messenger RNA and the amino acid sequence of epimerase demonstrates that the first 26 bases in the mRNA are transcribed but do not take part in translation of epimerase.     

Protease secretion in Neurospora crassa

Secreted and constitutive intracellular proteases of $\text{Neurospora crassa}$ differ with regard to inhibitor sensitivity, substrate specificity, isoelectric points and other properties. Upon the induction of protease secretion the enzymes released from the mycelium are formed $\text{de novo}$ as demonstrated by density labelling with D₂O. Vesicles which contain the constitutive intracellular proteases are, therefore, not involved in the secretion of proteases.     

Imidazole acetic acid as a substitute for cAMP.

The ability of imidazole acetic acid (IA) to substitute for cAMP was demonstrated by use of a series of strains carrying a lesion in the $\text{cya}$ structural gene. The substitution of IA for cAMP was specific for the L-arabinose operon in that this compound was ineffective in substituting for cAMP in the lactose or maltose catabolic systems. The cAMP receptor protein (CRP) and the $\text{araC}$ gene product were necessary for the IA mediated induction of the L-arabinose system.     

The amino terminal sequence of ATP-phosphoribosyltransferase, the first gene product of the histidine operon.

The amino terminal sequence of ATP-phosphoribosyltransferase of $\text{Salmonella typhimurium}$ has been determined by automated Edman degradation. Since this protein is the first gene product of the histidine operon, comparison of its amino terminal sequence with the genetic code allows the deduction of a partial base sequence of its mRNA and the corresponding DNA. Five of the first nine residues of ATP-phosphoribosyltransferase align with the amino terminal sequence of histidinol dehydrogenase, the second gene product of the histidine operon.     

The role of molybdenum in formation of the NADPH-nitrate reductase by Aspergillus nidulans

Non-nitrate reducing mutants of $\text{Aspergillus}$$\text{nidulans}$ have been noted to produce either a nitrate inducible or constitutive NADPH-cytochrome $\text{c}$ reductase which resides in either a 4.5s or a 7.8s protein. The latter closely resembles the nitrate inducible, FAD dependent NADPH-nitrate reductase from the wild type. Measurement of flavin adenine dinucleotide (FAD) and molybdenum (Mo) in these two proteins revealed significant differences particularly in Mo. The concepts that a nitrate inducible $\text{nia}$ gene product constitutes the major flavin bearing component of the enzyme and that a constitutively produced $\text{cnx}$ gene product is implicated in formation of the larger Mo bearing multimer are further supported.     

Crystallography of Bacillus subtilis levansucrase

Levansucrase, an exocellular enzyme, has been isolated from a high producer mutant, the BS5C4 constitutive strain, of Bacillus subtilis. Three crystalline forms have been obtained, all three belonging to the orthorhombic space group P2₁2₁2₁. The most suitable form for a three-dimensional structure investigation has cell dimensions, $\text{a = 68}\text{A}\text{?}$, $\text{b = 125}\text{A}\text{?}$, $\text{c = 54}\text{A}\text{?}$. There is one molecule in the asymmetric unit.     

Synthesis of the E. coli pyruvate dehydrogenase complex: non-dependence on 3'-5'-cyclic AMP

A determination of the level of the pyruvate dehydrogenase complex in a 3′–5′-c-AMP deficient mutant of $\text{E.}$$\text{coli}$ K12 has been carried out. The deficiency has no effect on specific activities for derivatives carrying either the inducible genes for two components of the complex or constitutive mutants. We conclude that synthesis of the complex is not sensitive to catabolite repression.     

On the origin of telocentric chromosomes in mammals

The origin of mammalian telocentric chromosomes is considered under the classical (fusion) and fission hypotheses using both theoretical analyses of the mechanisms proposed under the two hypotheses, and the published chromosomal data for 723 mammal species. Telocentrics are defined on the basis of short arm size (S_w) as chromosomes with S_w < 0·1(Imai, 1976). The fusion hypothesis lacks adequate models for producing these telocentrics, but their origin is readily understood under the fission hypothesis. Based on these analyses, I propose a cyclical model of chromosome change, symbolized:

in which T, A, and $\text{M}$ are, respectively, telocentric, acrocentric, and meta-, submeta- and subtelocentric chromosomes. The chief elements of this model are centric fission $\text{(}\text{M}\text{→ T + T)}$, tandem growth of constitutive heterochromatin (T → A), and pericentric inversion $\text{(A →}\text{M}\text{)}$. Under this model, therefore, mammalian karyotypes have an overall tendency, with occasional reversals, to evolve higher numbers of both chromosomes and chromosome arms.