Protein mimicry of DNA from crystal structures of the uracil-DNA glycosylase inhibitor protein and its complex with Escherichia coli uracil-DNA glycosylase 1 1Edited by D. C. Rees

Putnam, Christopher D.; Shroyer, Mary Jane N.; Lundquist, Amy J.; Mol, Clifford D.; Arvai, A.S.; Mosbaugh, Dale W.; Tainer, John A.

doi:10.1006/jmbi.1999.2605

Cited by 121 publications

(172 citation statements)

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“…Comparison of amino acid sequence of E. coli and mycobacterial UDGs Also shown are the residues at equivalent positions in M. tuberculosis, M. smegmatis, human and HSV UDGs [adapted from Putnam et al (1999) and Ravishankar et al (1998)]. …”

Section: Resultsmentioning

confidence: 99%

“…Several X-ray crystal structures of Ugi and its complexes with UDGs have elegantly demonstrated the structural features involved in their interaction (Savva & Pearl, 1995;Mol et al, 1995;Ravishankar et al, 1998;Putnam et al, 1999). The EcoUDG-Ugi interface is defined by two sets of predominant interactions: (i) an interaction between the hydrophobic cavity of Ugi (formed between the a2 helix and the b sheet) and L191 of the DNA intercalation loop of UDG, and (ii) the interactions of the b1-edge of Ugi in the DNA-binding groove of UDG via direct or the watermediated hydrogen bonds (Ravishankar et al, 1998;Putnam et al, 1999). Other studies (Lundquist et al, 1997; have shown that mutations in Ugi which compromise or abolish either the hydrophobic cavity interaction (M24K) or the b1-edge interaction (S21P, L23R, E20A) result in formation of complexes with EcoUDG that are reversible, suggesting that both classes of interaction are necessary to form a 'locked' complex.…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, a stopped flow kinetic study suggested a two-step ('docking' and 'locking') mechanism of EcoUDG-Ugi complex formation (Bennett et al, 1993). The co-crystal structures of various UDGs with Ugi have been resolved and show an extraordinary conservation in their overall architecture (Savva & Pearl, 1995;Mol et al, 1995;Ravishankar et al, 1998;Putnam et al, 1999). Interestingly, these studies have identified Ugi as a transition-state substrate mimic, making it an attractive model system to understand the mechanistic aspects of protein-protein interaction.…”

Section: Introductionmentioning

confidence: 99%

“…Interestingly, these studies have identified Ugi as a transition-state substrate mimic, making it an attractive model system to understand the mechanistic aspects of protein-protein interaction. Ugi primarily uses two of its structural elements, the hydrophobic pocket between the a2 helix and the antiparallel b sheet, and the unusually shaped b1-edge (beginning from the N-terminal side of Q19 to just past E28; Putnam et al, 1999) to form an exceptionally stable complex with UDGs. The hydrophobic pocket houses the L191 side chain of the DNA intercalation loop ) and the b1-edge interacts with the key active-site residues such as the water-activating loop, 63-QDPYH-67, the Pro-Ser loop; 84-AIPPS-88 and the DNA intercalation loop of EcoUDG (Putnam et al, 1999).…”

Section: Introductionmentioning

confidence: 99%

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Complexes of the uracil-DNA glycosylase inhibitor protein, Ugi, with Mycobacterium smegmatis and Mycobacterium tuberculosis uracil-DNA glycosylases

2003

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Uracil, a promutagenic base, appears in DNA either by deamination of cytosine or by incorporation of dUMP by DNA polymerases. This unconventional base in DNA is removed by uracil-DNA glycosylase (UDG). Interestingly, a bacteriophage-encoded short polypeptide, UDG inhibitor (Ugi), specifically inhibits UDGs by forming a tight complex. Three-dimensional structures of the complexes of Ugi with UDGs from Escherichia coli, human and herpes simplex virus have shown that two of the structural elements in Ugi, the hydrophobic pocket and the β1-edge, establish key interactions with UDGs. In this report the characterization of complexes of Ugi with UDGs from Mycobacterium tuberculosis, a pathogenic bacterium, and Mycobacterium smegmatis, a widely used model organism for the former, is described. Unlike the E. coli (Eco) UDG-Ugi complex, which is stable to treatment with 8 M urea, the mycobacterial UDG-Ugi complexes dissociate in 5–6 M urea. Furthermore, the Ugi from the complexes of mycobacterial UDGs can be exchanged by the DNA substrate. Interestingly, while EcoUDG sequestered Ugi into the EcoUDG-Ugi complex when incubated with mycobacterial UDG-Ugi complexes, even a large excess of mycobacterial UDGs failed to sequester Ugi from the EcoUDG-Ugi complex. However, the M. tuberculosis (Mtu) UDG-Ugi complex was seen when MtuUDG was incubated with M. smegmatis (Msm) UDG-Ugi or EcoUDG(L191G)-Ugi complexes. The reversible nature of the complexes of Ugi with mycobacterial UDGs (which naturally lack some of the structural elements important for interaction with the β1-edge of Ugi) and with mutants of EcoUDG (which are deficient in interaction with the hydrophobic pocket of Ugi) highlights the significance of both classes of interaction in formation of UDG-Ugi complexes. Furthermore, it is shown that even though mycobacterial UDG-Ugi complexes dissociate in 5–6 M urea, Ugi is still a potent inhibitor of UDG activity in mycobacteria.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Complexes of the uracil-DNA glycosylase inhibitor protein, Ugi, with Mycobacterium smegmatis and Mycobacterium tuberculosis uracil-DNA glycosylases

2003

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“…Pathway progession may also be regulated through the mimicry of DNA or protein partners. There are several examples of proteins mimicking DNA interactions to form complexes (Putnam and Tainer, 2005) and these were initially identified in the BER pathway (Mol et al, 1995,Savva and Pearl, 1995,Putnam et al, 1999. A significant example of protein interface mimicry occurs in Rad51 filament formation, which is central to HRR steps.…”

Section: Perspectivesmentioning

confidence: 99%

Developing master keys to brain pathology, cancer and aging from the structural biology of proteins controlling reactive oxygen species and DNA repair

2007

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This review is focused on proteins with key roles in pathways controlling either reactive oxygen species or DNA damage responses, both of which are essential for preserving the nervous system. An imbalance of reactive oxygen species or inappropriate DNA damage response likely causes mutational or cytotoxic outcomes, which may lead to cancer and/or aging phenotypes. Moreover, individuals with hereditary disorders in proteins of these cellular pathways have significant neurological abnormalities. Mutations in a superoxide dismutase, which removes oxygen free radicals, may cause the neurodegenerative disease amyotrophic lateral sclerosis. Additionally, DNA repair disorders that affect the brain to varying extents include ataxia-telangiectasia-like disorder, Cockayne syndrome or Werner syndrome. Here, we highlight recent advances gained through structural biochemistry studies on enzymes linked to these disorders and other related enzymes acting within the same cellular pathways. We describe the current understanding of how these vital proteins coordinate chemical steps and integrate cellular signaling and response events. Significantly, these structural studies may provide a set of master keys to developing a unified understanding of the survival mechanisms utilized after insults by reactive oxygen species and genotoxic agents, and also provide a basis for developing an informed intervention in brain tumor and neurodegenerative disease progression.

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Mass spectrometry‐based analysis of macromolecular complexes ofStaphylococcus aureusuracil‐DNA glycosylase and its inhibitor reveals specific variations due to naturally occurring mutations

et al. 2019

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The base excision repair pathway plays an important role in correcting damage induced by either physiological or external effects. This repair pathway removes incorrect bases from the DNA . The uracil base is among the most frequently occurring erroneous bases in DNA , and is cut out from the phosphodiester backbone via the catalytic action of uracil‐ DNA glycosylase. Uracil excision repair is an evolutionarily highly conserved pathway and can be specifically inhibited by a protein inhibitor of uracil‐ DNA glycosylase. Interestingly, both uracil‐ DNA glycosylase ( Staphylococcus aureus uracil‐ DNA glycosylase; SAUDG ) and its inhibitor ( S. aureus uracil‐ DNA glycosylase inhibitor; SAUGI ) are present in the staphylococcal cell. The interaction of these two proteins effectively decreases the efficiency of uracil‐ DNA excision repair. The physiological relevance of this complexation has not yet been addressed in detailed; however, numerous mutations have been identified within SAUGI . Here, we investigated whether these mutations drastically perturb the interaction with SAUDG . To perform quantitative analysis of the macromolecular interactions, we applied native mass spectrometry and demonstrated that this is a highly efficient and specific method for determination of dissociation constants. Our results indicate that several naturally occurring mutations of SAUGI do indeed lead to appreciable changes in the dissociation constants for complex formation. However, all of these K d values remain in the nanomolar range and therefore the association of these two proteins is preserved. We conclude that complexation is most likely preserved even with the naturally occurring mutant uracil‐ DNA glycosylase inhibitor proteins.

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Protein mimicry of DNA from crystal structures of the uracil-DNA glycosylase inhibitor protein and its complex with Escherichia coli uracil-DNA glycosylase 1 1Edited by D. C. Rees

Cited by 121 publications

References 50 publications

Complexes of the uracil-DNA glycosylase inhibitor protein, Ugi, with Mycobacterium smegmatis and Mycobacterium tuberculosis uracil-DNA glycosylases

Complexes of the uracil-DNA glycosylase inhibitor protein, Ugi, with Mycobacterium smegmatis and Mycobacterium tuberculosis uracil-DNA glycosylases

Developing master keys to brain pathology, cancer and aging from the structural biology of proteins controlling reactive oxygen species and DNA repair

Mass spectrometry‐based analysis of macromolecular complexes ofStaphylococcus aureusuracil‐DNA glycosylase and its inhibitor reveals specific variations due to naturally occurring mutations

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