Limited proteolysis and site-directed mutagenesis of the NF-κB p50 DNA-binding subunit

Sodeoka, Mikiko; Larson, Christopher J.; Chen, Lin; Lane, William S.; Verdine, Gregory L.

doi:10.1016/s0960-894x(00)80294-2

Cited by 40 publications

(5 citation statements)

References 9 publications

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“…The position of glutathione S-transferase (Jaffray et al, 1995). p50 RHR-c (residues the second RHRn p65 domain was determined to be in an orientation 243-366) was purified by a procedure similar to that used for the in the complex similar to the first RHRn, but with fewer contacts to RHR of p50 (Sodeoka et al, 1993a). The p65 RHR (residues 1-323) other domains, and is thus not well ordered.…”

Section: Associating Protein (Ib␣; Karyopherin-␣) That Imposesmentioning

confidence: 99%

Structure of an IκBα/NF-κB Complex

Jacobs

Harrison

1998

Cell

775

723

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The inhibitory protein, IkappaBalpha, sequesters the transcription factor, NF-kappaB, as an inactive complex in the cytoplasm. The structure of the IkappaBalpha ankyrin repeat domain, bound to a partially truncated NF-kappaB heterodimer (p50/ p65), has been determined by X-ray crystallography at 2.7 A resolution. It shows a stack of six IkappaBalpha ankyrin repeats facing the C-terminal domains of the NF-kappaB Rel homology regions. Contacts occur in discontinuous patches, suggesting a combinatorial quality for ankyrin repeat specificity. The first two repeats cover an alpha helically ordered segment containing the p65 nuclear localization signal. The position of the sixth ankyrin repeat shows that full-length IkappaBalpha will occlude the NF-kappaB DNA-binding cleft. The orientation of IkappaBalpha in the complex places its N- and C-terminal regions in appropriate locations for their known regulatory functions.

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Section: Associating Protein (Ib␣; Karyopherin-␣) That Imposesmentioning

confidence: 99%

Structure of an IκBα/NF-κB Complex

Jacobs

Harrison

1998

Cell

775

723

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“…The PdxA and PdxJ genes were subcloned from the Kohara phage E. coli DNA library . PCR was used to amplify each gene while simultaneously introducing a ribosome binding site and restriction sites before ligation into the pLM1 expression vector the overexpressed target proteins were purified to homogeneity using a three-step procedure involving hydroxyapatite, anion exchange, and gel filtration columns.…”

mentioning

confidence: 99%

Biosynthesis of Vitamin B₆: The Oxidation of 4-(Phosphohydroxy)-l-threonine by PdxA

et al. 1998

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“…The amounts (pmol) of identified amino acids and background (in parentheses) at each cycle were 282.4 (0), 56.5 (13.4), 33.0 (1.6), 32.5 (3.6), 23.3 (5.9), 21.8 (7.5), 0 (0), 15 (Fig. 3).…”

mentioning

confidence: 99%

Evidence for a non-alpha-helical DNA-binding motif in the Rel homology region.

Liu

Sodeoka

Lane

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

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The Fig. 1), when labeled at multiple sites with 5-bromo-2'-deoxyuridine (BrdUrd), underwent photocrosslinking to NF-KB, the p65 homodimer, and the p50 homodimer. In the present studies, we utilized 3-IFN oligonucleotides containing BrdUrd at single positions within or flanking the NF-KB element to identify individual positions within the 3-IFN site that underwent efficient crosslinking and to locate amino acid residues ofp50 which are near these base pairs in the protein-DNA interface.Here we report the direct identification of two amino acids which are crosslinked to BrdUrd-substituted f3-IFN oligonucleotides and the importance of these amino acids in DNAbinding properties ofp50. Secondary structure analysis ofthe stretch of peptide which was crosslinked to DNA revealed a plausible DNA-binding motifin p50: (3-strand-turn-1-strand.MATERIALS AND METHODS Materials. BrdUrd phosphoramidite was from Peninsula Laboratories. Oligonucleotides were synthesized on an Applied Biosystems 380A DNA synthesizer. The UV transilluminator (model 3-3000, four 15-W 300-nm bulbs, 21 cm x 38 cm UV filter glass) was from Fotodyne (New Berlin, WI *To whom reprint requests should be addressed. 908The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement"in accordance with 18 U.S.C. §1734 solely to indicate this fact.

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Limited proteolysis and site-directed mutagenesis of the NF-κB p50 DNA-binding subunit

Cited by 40 publications

References 9 publications

Structure of an IκBα/NF-κB Complex

Structure of an IκBα/NF-κB Complex

Biosynthesis of Vitamin B₆: The Oxidation of 4-(Phosphohydroxy)-l-threonine by PdxA

Evidence for a non-alpha-helical DNA-binding motif in the Rel homology region.

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Limited proteolysis and site-directed mutagenesis of the NF-κB p50 DNA-binding subunit

Cited by 40 publications

References 9 publications

Structure of an IκBα/NF-κB Complex

Structure of an IκBα/NF-κB Complex

Biosynthesis of Vitamin B6: The Oxidation of 4-(Phosphohydroxy)-l-threonine by PdxA

Evidence for a non-alpha-helical DNA-binding motif in the Rel homology region.

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Biosynthesis of Vitamin B₆: The Oxidation of 4-(Phosphohydroxy)-l-threonine by PdxA