Breaking open a protein barrel

Kallenbach, Neville R.

doi:10.1073/pnas.071051798

Cited by 9 publications

(19 citation statements)

References 32 publications

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“…The space group of BPTI-20st and BPTI- [5,55]st was P2 1 2 1 2 1 and P12 1 1 with four and two molecules per unit cell, respectively. High-resolution anisotropic models of BPTI-20st and BPTI- [5,55]st were determined at resolutions of 1.39 and 1.09 Å, respectively, and with crystallographic R factors of 0.1565 and 0.1811 (Table 1). All residue side chains had excellent electron density maps, in both BPTI-20st and BPTI- [5,55]st.…”

Section: Resultsmentioning

confidence: 99%

“…The effects of multiple alanine substitutions were surprisingly few. The overall structure of BPTI-20st was very similar to those of BPTI- [5,55]st and the wild-type BPTI, and the side-chain configurations of the deeply buried residues were almost perfectly retained. Alternative or new side-chain configurations were observed for a few partially buried residues.…”

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confidence: 79%

“…BPTI-21 (containing 21 alanines) was first designed by simultaneously substituting the least destabilizing BPTI- [5,55] residues to alanines (6), according to an alanine scanning experiment, in which each residue was individually changed to alanine and the stability of the mutated BPTIs were experimentally measured (17). Among the substitution sites, three residues are fully buried and two are partially buried.…”

mentioning

confidence: 99%

“…In the present study, we stabilized BPTI-21 by introducing an A14G substitution (18) that yielded BPTI-20st. As a control for the native structure, we determined the structure of a stabilized BPTI- [5,55] variant 55]st; BPTI- [5,55] with the A14G mutation), which is a single-disulfide-bonded BPTI variant regarded as a reference native molecule in several protein folding experiments (19). The effects of multiple alanine substitutions were surprisingly few.…”

mentioning

confidence: 99%

“…Indeed, an astronomical number of sequences are generated by the combination of the 20 natural amino acids at each site of a sequence, but few of these can fold into native, fully functional protein structures. However, the information contained in natural amino acid sequences is redundant (2,3); namely, a small fraction of the information can fully determine a protein structure (4,5). Protein sequence simplification has emerged as a method of choice in deciphering the determinants of protein structures.…”

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confidence: 99%

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Crystal structure of an extensively simplified variant of bovine pancreatic trypsin inhibitor in which over one-third of the residues are alanines

Islam¹,

Sohya²,

Noguchi

et al. 2008

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

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We report the high-resolution crystal structures of an extensively simplified variant of bovine pancreatic trypsin inhibitor containing 20 alanines (BPTI-20st) and a reference single-disulfide-bonded variant (BPTI-[5,55]st) at, respectively, 1.39 and 1.09 Å resolutions. The sequence was simplified based on the results of an alanine scanning experiment, as reported previously. The effects of the multiple alanine substitutions on the overall backbone structure were surprisingly small (C ␣ atom RMSD of 0.53 Å) being limited to small local structural perturbations. Both BPTI variants retained a wild-type level of trypsin inhibitory activity. The side-chain configurations of residues buried in the hydrophobic cores (<30% accessible surface area) were almost perfectly retained in both BPTI-20st and BPTI-[5,55]st, indicating that neither multiple alanine replacements nor the removal of the disulfide bonds affected their precise placements. However, the side chains of three partially buried residues (Q31, R20, and to some extent Y21) and several unburied residues rearranged into alternative dense-packing structures, suggesting some plasticity in their shape complementarity. These results indicate that a protein sequence simplified over its entire length can retain its densely packed, native side-chain structure, and suggest that both the design and fold recognition of natively folded proteins may be easier than previously thought.native structure ͉ protein design ͉ protein folding ͉ protein structure ͉ sequence simplification T he protein folding problem (1), or how a protein structure is encoded in its amino acid sequence, remains fundamentally unsolved. Indeed, an astronomical number of sequences are generated by the combination of the 20 natural amino acids at each site of a sequence, but few of these can fold into native, fully functional protein structures. However, the information contained in natural amino acid sequences is redundant (2, 3); namely, a small fraction of the information can fully determine a protein structure (4, 5). Protein sequence simplification has emerged as a method of choice in deciphering the determinants of protein structures.Information redundancy can be conceptually classified into class and site redundancy (6). Class redundancy refers to the overlapping physicochemical properties of the amino acids, and it can be removed by designing proteins with a reduced amino acid alphabet (7). Class redundancy has been reported in the Src SH3 domain, for which a functional variant with 70% of its sequence encoded by only five amino acids types was identified by phage display (8). Similarly, simplified variants of chorismate mutase (9) and orotate phosphoribosyl transferase (10), with sequences predominantly encoded by only nine amino acid types, rescued auxotrophic host cells lacking the respective enzymes.Site redundancy refers to the occurrence of several residues in a protein's amino acid sequence that do not contribute to the formation of the structure, except that their backbone atoms act as l...

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Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 79%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Crystal structure of an extensively simplified variant of bovine pancreatic trypsin inhibitor in which over one-third of the residues are alanines

Islam¹,

Sohya²,

Noguchi

et al. 2008

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

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Catalytic residues in hydrolases: analysis of methods designed for ligand-binding site prediction

Prymula

Jadczyk

Roterman

2010

J Comput Aided Mol Des

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The comparison of eight tools applicable to ligand-binding site prediction is presented. The methods examined cover three types of approaches: the geometrical (CASTp, PASS, Pocket-Finder), the physicochemical (Q-SiteFinder, FOD) and the knowledge-based (ConSurf, SuMo, WebFEATURE). The accuracy of predictions was measured in reference to the catalytic residues documented in the Catalytic Site Atlas. The test was performed on a set comprising selected chains of hydrolases. The results were analysed with regard to size, polarity, secondary structure, accessible solvent area of predicted sites as well as parameters commonly used in machine learning (F-measure, MCC). The relative accuracies of predictions are presented in the ROC space, allowing determination of the optimal methods by means of the ROC convex hull. Additionally the minimum expected cost analysis was performed. Both advantages and disadvantages of the eight methods are presented. Characterization of protein chains in respect to the level of difficulty in the active site prediction is introduced. The main reasons for failures are discussed. Overall, the best performance offers SuMo followed by FOD, while Pocket-Finder is the best method among the geometrical approaches.Electronic supplementary materialThe online version of this article (doi:10.1007/s10822-010-9402-0) contains supplementary material, which is available to authorized users.

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Thermodynamic Properties of BPTI Variants with Highly Simplified Amino Acid Sequences

Kato

Yamada

Nakamura

et al. 2007

Journal of Molecular Biology

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Breaking open a protein barrel

Cited by 9 publications

References 32 publications

Crystal structure of an extensively simplified variant of bovine pancreatic trypsin inhibitor in which over one-third of the residues are alanines

Crystal structure of an extensively simplified variant of bovine pancreatic trypsin inhibitor in which over one-third of the residues are alanines

Catalytic residues in hydrolases: analysis of methods designed for ligand-binding site prediction

Thermodynamic Properties of BPTI Variants with Highly Simplified Amino Acid Sequences

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