Structural features of normal and complemented forms of the Neurospora isopropylmalate isomerase

Reichenbecher, V E; Gross, S. R.

doi:10.1128/jb.133.2.802-810.1978

Cited by 10 publications

(7 citation statements)

References 26 publications

(15 reference statements)

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“…The enzyme is significantly less stable at pH 6, in contrast to the enzyme from N. crassa, which was found to be more stable at pH 6 (10). In addition, we found no stabilization of the S. typhimurium enzyme by glycerol, which was reported to stabilize the isopropylmalate isomerase isolated from yeasts and from N. crassa (2,25).…”

Section: Resultsmentioning

confidence: 65%

“…Since the leuC and leuD genes are cotranscribed and the expression of the leuA and leuB genes is not affected in leuC and leuD mutant strains, the suggestion of Bigelis and Umbarger (2) that one ofthe polypeptides has a regulatory function affecting the leucine operon can be eliminated. The finding that the isopropylmalate isomerase purified from N. crassa, which is a single polypeptide of 90,000 daltons, is nonrandomly cleaved into two polypeptide fragments of 56,000 and 37,000 daltons (25) suggests that the gene coding for this enzyme might be the result of a fusion of two genes during the evolution of this species. Thus, the native form of the N. crassa isopropylmalate isomerase could contain two distinct domains in its tertiary structure which represent the two ancestral polypeptides that are now joined in a region which is a preferential site for peptide cleavage.…”

Section: Discussionmentioning

confidence: 99%

“…The complexity ofthe suppression pattern of different leuD mutations by the same supQ mutation is most satisfactorily explained if the leuC-leuD region of the chromosome is composed of two separate genes (8,(13)(14)(15). Whether the leuC-leuD region in E. coli and S. typhimurium actually codes for two different polypeptides has recently been questioned, primarily because the purification of the isopropylmalate isomerase from yeasts and Neurospora crassa has shown that in both of these species this enzyme is composed of only one polypeptide with a molecular weight of 90,000 (1,25). In vivo complementation data for the N. crassa isopropylmalate isomerase had indicated t Present address: Department of Cell Biology, University of Texas Health Science Center at Dallas, Dallas, TX 75235. the involvement of two separate genes (in different linkage groups) (11), suggesting that the isopropylmalate isomerase in this organism was a complex enzyme composed of two different polypeptide chains; however, one gene has now been shown to be the structural gene for the enzyme, and the other is involved in regulation (12,24).…”

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confidence: 99%

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Wild-type isopropylmalate isomerase in Salmonella typhimurium is composed of two different subunits

Fultz¹,

Kemper²

1981

J Bacteriol

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The isopropylxnalate isomerase in Salmonella typhimurium is the second enzyme specific for leucine biosynthesis. It is a complex enzyme composed of two subunits which are coded for by two genes of the leucine operon, leuC and leuD. The two polypeptides have been shown to copurify through successive ammonium sulfate fractionations and have been identified on sodium dodecyl sulfate-polyacrylamide gels as having molecular weights of 51,000 (leuC gene product) and 23,500 (leuD gene product). They have also been shown to be fairly stable, since in vitro complementation of cell-free extracts of leuC and leuD mutant strains was demonstrated, with only a 40% loss of activity 16 h after preparation of the extracts. The native isopropylmalate isomerase was shown to have a Km for its substrate a-isopropyhnalate of 3 x 10-4 M. flr-19 leuD657 A(leuD-ara) 798 A(gpt-proBA-supQ-newD)47 fol-101IF' leu ara

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Section: Resultsmentioning

confidence: 65%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Wild-type isopropylmalate isomerase in Salmonella typhimurium is composed of two different subunits

Fultz¹,

Kemper²

1981

J Bacteriol

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“…Similar gene fusion has been suggested to explain the differences in the tryptophan operons of bacterial and eucaryotic species (5). Supportive evidence is given by the fact that the N. crassa isopropylmalate isomerase is preferentially cleaved into two major fragments of approximately 56,000 and 37,000 daltons (20).…”

Section: Discussionmentioning

confidence: 99%

Salmonella typhimurium newD and Escherichia coli leuC genes code for a functional isopropylmalate isomerase in Salmonella typhimurium-Escherichia coli hybrids

Fultz¹,

Kwoh²,

Kemper³

1979

J Bacteriol

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The supQ newD gene substitution system in Salmonella typhimurium restores leucine prototrophy to leuD mutants by providing the newD gene product which is capable of replacing the missing leuD polypeptide in the isopropylmalate isomerase, a complex of the leuC and leuD gene product. Mutations in the supQ gene are required to make the newD protein available. An Escherichia coli F' factor was constructed which carried supQ- newD+ from S. typhimurium on a P22-specialized transducing genome. This F' pro lac (P22dsupQ394newD) episome was transferred into S. typhimurium strains containing th leuD798-ara deletion; the resulting merodiploid strains had a Leu+ phenotype, indicating that supQ- newD+ is dominant over supQ+ newD+, and eliminating the possibility that the supQ gene codes for a repressor of the newD gene. Furthermore, transfer of the F' pro lac (P22dsupQ39newD) into E. coli leuD deletion strains restored leucine prototrophy, showing that the S. typhimurium newD gene can complment the E. coli leuC gene. Growth rates of the S. typhimurium-E coli hybrid strains indicated that the mutant isopropylmalate isomerase in these strains does not induce a leucine limitation, as it does in S. typhimurium leuD supQ mutants. In vitro activity of the mutant isopropylmalate isomerase was demonstrated; the Km values for alpha-isopropylmalate of both the S. typhimurium leuC-newD isomerase and the S. typhimurium-E. coli hybrid isomerase were as much as 100 times higher than the Km values for alpha-isopropylmalate of the wild-type enzyme, which was 3 x 10(-4) M. Mutagenesis of E. coli leuD deletion strains failed to restore leucine prototrophy, indicating that E. coli does not have genes analogous to the S. typhimurium supQ newD genes, of that, if present, activation of a newD is a rare event or is lethal to the cell.

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“…Able to use a-isopropylmalate to support growth of leu4 mutants and for induction of aisopropylmalate isomerase and P-isopropylmalate dehydrogenase, in contrast to ipm+ strains, which are unable to take up this intermediate (870,871). ipm-2: isopropyhnalate permeation-2 Unmapped.…”

Section: In(il -) Ir)h4250mentioning

confidence: 99%

Chromosomal loci of Neurospora crassa

Perkins¹,

Radford²,

Newmeyer³

et al. 1982

Microbiol Rev

259

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I I ' ~~~~T h Ñ CiOr(r r N z ADDITIONAL LOCI IN LINKAGE GROUP ILinked to mt (8 to 12%) Linked to mt, acr-3 (5%) Right of arg-i (5/29) Linked to so and aro-8 Right of un-18 Between ad-5 and the centromere Between arg-i (4%) and his-3 (I to 2%) Between In(H4250) and T(39311) Left of mt (23%) Linked to mt (2%) and aza-i (39%o) Left of nit-2 (30%) Left of mt (14%) Linked to un-i (9%), probably to the right Linked to hom (1%), probably to the right Linked to mt (17 to 22%) Between T(NM103) and al-2 (18%) Between T(4540) and cr-2Between thi-i (12 to 20%/6) and ad-9 (5%) Linked to ad-5 (1/54), probably to the left Linked to mt (6%) Linked to cys-5 (<1%), probably to the right Right of ad-9 (12%), linked to al (0/76) Right of his-3 (2%) Between the T(AR173) breakpoints Linked to al-2 (10%) and nic-i (1 to 5%) Linked to ad-9 (0/44); right of thi-i (2%) Linked to mt (5%), probably to the left Linked to al-I (10%) Right of os-i (3%) Between nit,i (5 to 19%) and al-I (6 to 19%io) Linked near arg-i Left of al-2 (25%o) Left of mt (16%) Right of nic-2 (6%) Linked to ad-9 (2%), probably to the right Left of cr-i (15%) Left of mt (10%) Probably between his-2 (3%) and cr-i (3%) Between T(NM103) and the right tip Between mt (20%) and arg-i (1%) Between ad-3 (6%) and al-i (16%) Between his-3 and nic-2 Between cyh-i (5%) and al-I (7%) Linked to cyh-i (18%), al-i (2%), and mb-2 (6%) Between arg-i (3%) and T(39311) Between tre (41%) and ad-9 Linked to mt (9%) Linked to mt (20%) Complex phenotype. Alternative requirements: 2-acetylaminofluorine, certain azo dyes, or certain single amino acids. Cold sensitive. Originated among progeny of a rib-i strain that had become tolerant to 2-acetylaminofluorine.

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Structural features of normal and complemented forms of the Neurospora isopropylmalate isomerase

Cited by 10 publications

References 26 publications

Wild-type isopropylmalate isomerase in Salmonella typhimurium is composed of two different subunits

Wild-type isopropylmalate isomerase in Salmonella typhimurium is composed of two different subunits

Salmonella typhimurium newD and Escherichia coli leuC genes code for a functional isopropylmalate isomerase in Salmonella typhimurium-Escherichia coli hybrids

Chromosomal loci of Neurospora crassa

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