scholarly journals RHO AND RIBOSOME MUTATION INTERACTION: LETHALITY OF rho-15 IN rpsL or rpsE STRAINS, AND rho-15 METHIONINE AUXOTROPHY IN rps  + STRAINS OF ESCHERICHIA COLI

Genetics ◽  
1979 ◽  
Vol 93 (2) ◽  
pp. 353-360
Author(s):  
S K Guterman ◽  
C L Howitt
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Growth Rate ◽  
Amino Acid ◽  
Genetic Background ◽  
Minimal Medium ◽  
Complex Medium ◽  
Single Amino Acid ◽  
Temperature Sensitive ◽  
K 12

ABSTRACT The phenotype of Escherichia coli K-12 carrying rho-15 in the genetic background DW319 ilv lacZ: :IS1 is described. Seventy-eight percent (70/90) of Ilv+ transductants acquired the following phenotype: temperature-sensitive growth on minimal salts medium, Ts+ growth on complex medium and suppression of the lac polar mutation. At 42° on minimal medium, the rho-15 transductants were cross-fed by a substance diffusing from Rho+ transductants or controls. The requirement for this substance was satisfied by methionine or cystathionine, but not by any other single amino acid or combination of amino acids, by spermidine, or by mono- or divalent cationic salts.—Transduction of rho-15 into four other Ilv- recipients revealed two phenotypic patterns. Recipients with rpsL or rpsE ribosomes yielded rho-15 transductants that were Ts on all media, or Ts on minimal medium whether or not methionine was present. The effect of the ribosome on expression of rho15 was confirmed by transduction of appropriate rps alleles into DW319, followed by co-transduction of rho-15 with Ilv+. The growth rate of double rho-15 rpsL or rho-15 rpsE strains was severely reduced at 42° in comparison with strains carrying any of these single mutations. Models for rho and ribosome interaction are presented.

scholarly journals Structural and functional analyses of Saccharomyces cerevisiae wild-type and mutant RNA1 genes.

1989 ◽  
Vol 9 (7) ◽  
pp. 2989-2999 ◽  
Author(s):  
H M Traglia ◽  
N S Atkinson ◽  
A K Hopper
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Open Reading Frame ◽  
Single Amino Acid ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Reading Frame ◽  
Genomic Deletions ◽  
Single Amino Acid Change ◽  
Carboxy Terminal

The yeast gene RNA1 has been defined by the thermosensitive rna1-1 lesion. This lesion interferes with the processing and production of all major classes of RNA. Each class of RNA is affected at a distinct and presumably unrelated step. Furthermore, RNA does not appear to exit the nucleus. To investigate how the RNA1 gene product can pleiotropically affect disparate processes, we undertook a structural analysis of wild-type and mutant RNA1 genes. The wild-type gene was found to contain a 407-amino-acid open reading frame that encodes a hydrophilic protein. No clue regarding the function of the RNA1 protein was obtained by searching banks for similarity to other known gene products. Surprisingly, the rna1-1 lesion was found to code for two amino acid differences from wild type. We found that neither single-amino-acid change alone resulted in temperature sensitivity. The carboxy-terminal region of the RNA1 open reading frame contains a highly acidic domain extending from amino acids 334 to 400. We generated genomic deletions that removed C-terminal regions of this protein. Deletion of amino acids 397 to 407 did not appear to affect cell growth. Removal of amino acids 359 to 397, a region containing 24 acidic residues, caused temperature-sensitive growth. This allele, rna1-delta 359-397, defines a second conditional lesion of the RNA1 locus. We found that strains possessing the rna1-delta 359-397 allele did not show thermosensitive defects in pre-rRNA or pre-tRNA processing. Removal of amino acids 330 to 407 resulted in loss of viability.

fusB is an allele of nadD, encoding nicotinate mononucleotide adenylyltransferase in Escherichia coli

Microbiology ◽  
2003 ◽  
Vol 149 (9) ◽  
pp. 2427-2433 ◽  
Author(s):  
Martin Stancek ◽  
Leif A. Isaksson ◽  
Monica Rydén-Aulin
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Temperature Sensitivity ◽  
Minimal Medium ◽  
Synthetic Medium ◽  
Normal Growth ◽  
Temperature Sensitive ◽  
Permissive Temperature ◽  
Atp Binding Site ◽  
Pleiotropic Phenotype

Isolation of the temperature-sensitive Escherichia coli mutant 72c has been described previously. The mutant allele was named fusB and causes a pleiotropic phenotype, the most striking features of which, besides temperature sensitivity, are the inability to grow on synthetic medium and supersensitivity to trimethoprim, an antibiotic that inhibits the C1 metabolism. This work shows that the fusB mutation is a frameshift mutation in the nadD gene that encodes nicotinate mononucleotide adenylyltransferase. The frameshift leads to a change of the last 10 amino acids and an addition of 17 amino acids. This lesion, renamed nadD72, leads to very little NAD+ and NADPH synthesis at the permissive temperature and essentially no synthesis at the non-permissive temperature. As a comparison, a new mutation in the nadD gene, with an amino acid change in the ATP-binding site, has been isolated. Its NAD+ synthesis is decreased at 30 °C but the level is still sufficient to support normal growth. At 42 °C, NAD+ synthesis is reduced further, which leads to temperature sensitivity on minimal medium. This mutation was designated nadD74. Thus, a small decrease in NAD+ levels affects ability to grow on minimal medium at 42 °C, while a large decrease leads to a more pleiotropic phenotype.

Growth and Changes in Pool and Macromolecular Components of Schizosaccharomyces Pombe During the Cell Cycle

1971 ◽  
Vol 9 (3) ◽  
pp. 701-717
Author(s):  
NOWELL STEBBING
Keyword(s):  
Cell Cycle ◽  
Amino Acids ◽  
Amino Acid ◽  
Schizosaccharomyces Pombe ◽  
Minimal Medium ◽  
Linear Growth ◽  
Dry Weight ◽  
Total Cell ◽  
Complex Medium ◽  
Synchronous Cultures

Amino acids, nucleotide and carbohydrate material were found to account for 46% of the total dry weight of pool material in Schizosaccharomyces pombe growing in minimal medium. The composition of the amino acid pool was also determined by autoanalysis and was found to be unaltered during growth in 2 M sorbitol, indicating that pool amino acids are not important in the osmoregulation of the cell. Kinetic analysis of the amino acid pool using 14C-labelled amino acids showed that amino acids accumulated from the medium enter an ‘expandable’ pool distinct from the ‘internal’ pool which is maintained during growth on minimal medium. Total RNA, protein, pool amino acid and pool ‘nucleotide’ material were estimated in synchronous cultures grown in minimal medium. All these components appeared to accumulate in an exponential manner during the cell cycle. Direct estimation of total cellular dry weight and the total pool in synchronous cultures showed that total cell dry weight increased exponentially and the pool did not fluctuate during growth in minimal medium. This contrasts with previous work on single cells of S. pombe grown in complex medium which showed that the dry weight of the pool fluctuates during the cell cycle and total cell dry weight increased linearly. Linear growth of S. pombe in malt extract broth can be accounted for by the presence of the second (‘expandable’) pool of amino acids formed during growth in complex medium. The phenomenon of linear growth during the cell cycle is shown to occur generally only in cells growing in complex medium. The phenomenon is considered in relation to mechanisms for controlling the size of the pool during growth in complex media.

scholarly journals A Single Amino Acid Substitution Converts γ-Glutamyltranspeptidase to a Class IV Cephalosporin Acylase (Glutaryl-7-Aminocephalosporanic Acid Acylase)

2004 ◽  
Vol 70 (10) ◽  
pp. 6324-6328 ◽  
Author(s):  
Hideyuki Suzuki ◽  
Chinatsu Miwa ◽  
Sayaka Ishihara ◽  
Hidehiko Kumagai
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Amino Acid Substitution ◽  
Starting Material ◽  
Single Amino Acid Substitution ◽  
Single Amino Acid ◽  
Cephalosporin Acylase ◽  
K 12 ◽  
Aspartyl Residue ◽  
Class Iv

ABSTRACT The aspartyl residue at position 433 of γ-glutamyltranspeptidase of Escherichia coli K-12 was replaced by an asparaginyl residue. This substitution enabled γ-glutamyltranspeptidase to deacylate glutaryl-7-aminocephalosporanic acid, producing 7-aminocephalosporanic acid, which is a starting material for the synthesis of semisynthetic cephalosporins.

scholarly journals Morphological and physiological effects of a single amino acid substitution in the patatin-like phospholipase CapV in Escherichia coli

2020 ◽  
Author(s):  
Fengyang Li ◽  
Heike Bähre ◽  
Manfred Rohde ◽  
Ute Römling
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Biofilm Formation ◽  
Morphological Plasticity ◽  
Single Amino Acid ◽  
E Coli ◽  
Single Amino Acid Change ◽  
Response To Stress ◽  
K 12

AbstractIn rod-shaped bacteria morphological plasticity occurs in response to stress, which blocks cell division to promote filamentation. We demonstrate here that overexpression of the patatin-like phospholipase variant CapVQ329R but not CapV causes pronounced sulA-independent pyridoxine-inhibited cell filamentation and restriction of swimming and flagella production of Escherichia coli K-12 derivative MG1655. Mutational analyses of CapVQ329R indicated conserved amino acids in canonical patatin-like phospholipase A motifs, but not the nucleophilic serine to be required for the observed phenotypes. Furthermore, CapVQ329R alters rdar biofilm formation including expression of the biofilm activator CsgD. Moreover, commensal and pathogenic E. coli strains and Salmonella typhimurium also responded with cell filamentation and alteration in biofilm formation. In conclusion, this work identifies the CapV variant CapVQ329R as a pleiotropic regulator, emphasizes a scaffold function for patatin-like phospholipases and highlights the role of a single amino acid change for the evolution of protein functionality.

scholarly journals DsdX Is the Second d-Serine Transporter in Uropathogenic Escherichia coli Clinical Isolate CFT073

2006 ◽  
Vol 188 (18) ◽  
pp. 6622-6628 ◽  
Author(s):  
Andrew T. Anfora ◽  
Rodney A. Welch
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Carbon Source ◽  
Clinical Isolate ◽  
Sole Carbon Source ◽  
Minimal Medium ◽  
Amino Acid Sequence Similarity ◽  
Predicted Amino Acid Sequence ◽  
E Coli ◽  
K 12

ABSTRACTd-Serine is an amino acid present in mammalian urine that is inhibitory toEscherichia colistrains lacking a functionaldsdAgene. Counterintuitively, adsdAstrain ofE. coliclinical isolate CFT073 hypercolonizes the bladder and kidneys of mice relative to wild type during a coinfection in the murine model of urinary tract infection. We are interested in the mechanisms for uptake ofd-serine in CFT073.d-Serine entersE. coliK-12 via CycA, thed-alanine transporter andd-cycloserine sensitivity locus. CFT073cycAcan grow on minimal medium withd-serine as a sole carbon source. ThedsdXgene of thedsdCXAlocus is a likely candidate for an additionald-serine transporter based on its predicted amino acid sequence similarity to gluconate transporters. In minimal medium, CFT073dsdXcan grow ond-serine as a sole carbon source; however, CFT073dsdX cycAcannot. Additionally, CFT073dsdXA cycAis not sensitive to inhibitory concentrations ofd-serine during growth on glycerol andd-serine minimal medium.d-[14C]serine uptake experiments with CFT073dsdX cycAharboringdsdXorcycArecombinant plasmids confirm thatd-serine is able to enterE. colicells via CycA or DsdX. In whole-celld-[14C]serine uptake experiments, DsdX has an apparentKmof 58.75 μM and aVmaxof 75.96 nmol/min/mg, and CycA has an apparentKmof 82.40 μM and aVmaxof 58.90 nmol/min/mg. Onlyd-threonine marginally inhibits DsdX-mediatedd-serine transport, whereasd-alanine, glycine, andd-cycloserine inhibit CycA-mediatedd-serine transport. DsdX or CycA is sufficient to transport physiological quantities ofd-serine, but DsdX is ad-serine-specific permease.

scholarly journals Recombination-Dependent Growth in Exonuclease-Depleted recBC sbcBC Strains of Escherichia coli K-12

Genetics ◽  
1996 ◽  
Vol 143 (3) ◽  
pp. 1101-1114 ◽  
Author(s):  
Lizanne Ryder ◽  
Gary J Sharples ◽  
Robert G Lloyd
Keyword(s):  
Escherichia Coli ◽  
Growth Rate ◽  
Dna Repair ◽  
Cell Viability ◽  
Replication Fork ◽  
Temperature Sensitive ◽  
Chromosome Duplication ◽  
New Gene ◽  
Dependent Growth ◽  
K 12

Abstract Analysis of the aroLM-sbcCD interval of the Escherichia coli K-12 chromosome revealed a new gene (rdgC) encoding a function required for growth in recombination-deficient recBC sbcBC strains. Deletion of rdgC does not reduce viability, conjugational recombination, or DNA repair in ret  +, recA, recB, reCF or recJ mutants. However, it makes the growth of recBC sbcBC strains reliant on the RecA, RecF, and RuvC proteins and, to a large extent, on RuvAB. The recBC sbcBC ΔrdgC ruvAB construct forms colonies, but cell viability is reduced to <5%. A recBC sbcBC ΔrdgC derivative carrying the temperature-sensitive recA200 allele grows at 32° but not 42°. Multicopy rdgC  + plasmids reduce the growth rate of recBC sbcBC strains, while multicopy sbcC plasmids that reactivate SbcCD nuclease cannot be maintained without RdgC protein. The data presented are interpreted to suggest that exonuclease-depleted recBC sbcBC strains have difficulty removing the displaced arm of a collapsed replication fork and that this problem is compounded in the absence of RdgC. Recombination then becomes necessary to repair the fork and allow chromosome duplication to be completed. The possibility that RdgC is an exonuclease is discussed.

Suppression of the ssb-1 and ssb-113 mutations of Escherichia coli by a wild-type rep gene, NaCl, and glucose

1982 ◽  
Vol 152 (2) ◽  
pp. 572-583
Author(s):  
E S Tessman ◽  
P K Peterson
Keyword(s):  
Escherichia Coli ◽  
Temperature Sensitivity ◽  
Genetic Background ◽  
Growth Defect ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Strong Suppression ◽  
Uv Sensitivity ◽  
Ssb Proteins ◽  
K 12

The ssb-1 mutation confers severe temperature sensitivity and UV sensitivity on many strains of Escherichia coli K-12 and C, including strain C1412. However, ssb-1 confers only slight temperature sensitivity and slight UV sensitivity on strain C1a, suggesting that strain C1a contains extragenic suppressors of ssb-1. We found that introduction of the wild-type rep gene from C1a into strain C1412 ssb-1 gave strong suppression of temperature sensitivity and moderate suppression of UV sensitivity. Also, the C1a rep+ gene mildly suppressed the temperature sensitivity conferred by the ssb-113 mutation, formerly called lexC113. Suppression of the C1412 ssb-1 growth defect by C1a rep+ rendered the cells Gro- for phi X174. In contrast to the positive suppression of ssb-1 and ssb-113 by a wild-type rep gene, mutant rep alleles enhanced the severity of the ssb-1 defect, with several C1a ssb-1 double mutants being either more temperature sensitive or more UV sensitive than C1a ssb-1, depending on which mutant rep allele was used. As a control, the same rep alleles in combination with a dnaB mutation gave an allele-independent increase in temperature sensitivity. Our results on suppression of ssb-1 by rep and on the role of the genetic background in this suppression suggested that the rep and ssb proteins interact to form a subcomplex of the total DNA replication complex and that this subcomplex has some function in repair. The effects of NaCl and glucose on suppression of both the temperature sensitivity and the UV sensitivity conferred by ssb-1 and ssb-113 are described. The degree of suppression of temperature sensitivity by salt or glucose was dependent on the source of the wild-type rep allele, as well as on the genetic background.

Structural and functional analyses of Saccharomyces cerevisiae wild-type and mutant RNA1 genes

1989 ◽  
Vol 9 (7) ◽  
pp. 2989-2999
Author(s):  
H M Traglia ◽  
N S Atkinson ◽  
A K Hopper
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Open Reading Frame ◽  
Single Amino Acid ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Reading Frame ◽  
Genomic Deletions ◽  
Single Amino Acid Change ◽  
Carboxy Terminal

The yeast gene RNA1 has been defined by the thermosensitive rna1-1 lesion. This lesion interferes with the processing and production of all major classes of RNA. Each class of RNA is affected at a distinct and presumably unrelated step. Furthermore, RNA does not appear to exit the nucleus. To investigate how the RNA1 gene product can pleiotropically affect disparate processes, we undertook a structural analysis of wild-type and mutant RNA1 genes. The wild-type gene was found to contain a 407-amino-acid open reading frame that encodes a hydrophilic protein. No clue regarding the function of the RNA1 protein was obtained by searching banks for similarity to other known gene products. Surprisingly, the rna1-1 lesion was found to code for two amino acid differences from wild type. We found that neither single-amino-acid change alone resulted in temperature sensitivity. The carboxy-terminal region of the RNA1 open reading frame contains a highly acidic domain extending from amino acids 334 to 400. We generated genomic deletions that removed C-terminal regions of this protein. Deletion of amino acids 397 to 407 did not appear to affect cell growth. Removal of amino acids 359 to 397, a region containing 24 acidic residues, caused temperature-sensitive growth. This allele, rna1-delta 359-397, defines a second conditional lesion of the RNA1 locus. We found that strains possessing the rna1-delta 359-397 allele did not show thermosensitive defects in pre-rRNA or pre-tRNA processing. Removal of amino acids 330 to 407 resulted in loss of viability.

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