Mapping the Distribution of Conformational Information Throughout a Protein Sequence

Gebhard, Leopoldo Germán; Risso, Valeria A.; Santos, Javier; Ferreyra, Raúl G.; Noguera, Martín Ezequiel; Ermácora, Mario R.

doi:10.1016/j.jmb.2006.01.095

Cited by 23 publications

(25 citation statements)

References 56 publications

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“…The number and the intricacy of its geometric constraints indicate that the conformational integrity of most of the two domains is required for catalysis, and that ESBL can only be enzymically active if able to fold in the native state (see Ref. 22 for a detailed analysis of the structure-activity relationship for ESBL). For this reason, probing the enzymic activity of the ESBL variants permits evaluation of the degree of conformational perturbation of the native state introduced by the helix replacement.…”

Section: Enzymic Activitymentioning

confidence: 99%

“…On the other hand, the triple mutant shows only traces of activity. A more detailed analysis of the catalytic activity showed that all of the of the above mutants exhibit Michaelis-Menten behavior allowing calculation of k cat and K m (Table II) and comparison with the catalytic efficiencies of the unperturbed protein and with previously characterized mutants 22 lacking altogether the helices under study. The k cat /K m value for a 9 and a 10 ESBL places these variants in the category of efficient catalysts.…”

Section: Enzymic Activitymentioning

confidence: 99%

“…The triple mutant, even though a weak catalyst, accelerates the hydrolysis of the substrate by a factor of 500 compared with the uncatalyzed reaction. 22 Importantly, the catalytic efficiency of a 9,10,12 ESBL (log(k cat /K m )>1. 6 22 ); suggests that it is a genuine enzyme, which in turn implies that at least a small fraction of the molecules of this mutant is still able to fold in a native-like structure.…”

Section: Enzymic Activitymentioning

confidence: 99%

“…22 We will also show that the exercise can mimic the creation of a protoenzyme that might further evolve to an efficient catalyst crossing sequence family boundaries while preserving the essential features of the fold.…”

Section: Introductionmentioning

confidence: 96%

“…22 Yet, protein functions and folds evolve by sequence changes, and therefore there should be a way to reconcile all the above properties in a single folding theory. To that end, evolutionary studies and experiments aimed to test under controlled conditions the relationship between sequence and structure are in great demand.…”

Section: Introductionmentioning

confidence: 99%

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Re‐engineering a β‐lactamase using prototype peptides from a library of local structural motifs

2009

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B. licheniformis exo-small b-lactamase (ESBL) has a complex architecture with twelve a helices and a five-stranded beta sheet. We replaced, separately or simultaneously, three of the ESBL a helices with prototype amphiphatic helices from a catalog of secondary structure elements. Although the substitutes bear no sequence similarity to the originals and pertain to unrelated protein families, all the engineered ESBL variants were found able to fold in native like structures with in vitro and in vivo enzymic activity. The triple substituted variant resembles a primitive protein, with folding defects such as a strong tendency to oligomerization and very low stability; however it mimics a non homologous recombinant abandoning the family sequence space while preserving fold. The results test protein folding and evolution theories.

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Section: Enzymic Activitymentioning

confidence: 99%

Section: Enzymic Activitymentioning

confidence: 99%

Section: Enzymic Activitymentioning

confidence: 99%

Section: Introductionmentioning

confidence: 96%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Re‐engineering a β‐lactamase using prototype peptides from a library of local structural motifs

2009

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Circular Permutation of Proteins

Meister

Kanwar

Ostermeier

2008

Protein Engineering Handbook

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Tools to evaluate the conformation of protein products

Manta

Obal

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et al. 2011

Biotechnology Journal

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Production of recombinant proteins is a process intensively used in the research laboratory. In addition, the main biotechnology market products are recombinant proteins and monoclonal antibodies. The biological (and clinical) properties of the protein product strongly depend on the conformation of the polypeptide. Therefore, assessment of the correct conformation of the produced protein is crucial. There is no single method to assess every aspect of protein structure or function. Depending on the protein, the methods of choice vary. There are general methods to evaluate not only mass and primary sequence of the protein, but also higher-order structure. This review outlines the principal techniques for determining the conformation of a protein from structural (biophysical methods) to functional (in vitro binding assays) analyses.

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Mapping the Distribution of Conformational Information Throughout a Protein Sequence

Cited by 23 publications

References 56 publications

Re‐engineering a β‐lactamase using prototype peptides from a library of local structural motifs

Re‐engineering a β‐lactamase using prototype peptides from a library of local structural motifs

Circular Permutation of Proteins

Tools to evaluate the conformation of protein products

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