On the routine use of soft X-rays in macromolecular crystallography. Part IV. Efficient determination of anomalous substructures in biomacromolecules using longer X-ray wavelengths

Mueller-Dieckmann, Christoph; Panjikar, Santosh; Schmidt, Andrea; Mueller, Simone; Kuper, Jochen; Geerlof, Arie; Wilmanns, Matthias; Singh, Rajesh; Tucker, Paul A.; Weiss, M.S.

doi:10.1107/s0907444906055624

Cited by 90 publications

(90 citation statements)

References 33 publications

(46 reference statements)

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“…Thrombin crystallizes in the E form in the presence of a point mutation, R77aA, that prevents auto-proteolysis (35, 36), or the double mutation C191A/C220A that perturbs the primary specificity pocket (34). Wild-type murine thrombin crystallizes in the E form (64) and so do C2a (65, 66), neuropsin (67) and trypsin (68). Complement factor C1r crystallizes in the E form (60), but assumes a collapsed conformation in its zymogen form (60).…”

Section: Allostery In Trypsin-like Enzymes: the E* And E Formsmentioning

confidence: 99%

Allostery in trypsin-like proteases suggests new therapeutic strategies

Gohara¹,

Di²

2011

Trends in Biotechnology

111

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Trypsin-like proteases are a large family of enzymes responsible for digestion, blood coagulation, fibrinolysis, development, fertilization, apoptosis and immunity. A current paradigm posits that the irreversible transition from an inactive zymogen to the active protease form enables productive interaction with substrate and catalysis. Analysis of the entire structural database reveals two distinct conformations of the active site: one fully accessible to substrate (E) and the other occluded by the collapse of a specific segment (E*). The allosteric E*-E equilibrium provides a reversible mechanism for activity and regulation besides the irreversible zymogen to protease conversion and points to new therapeutic strategies aimed at inhibiting or activating the enzyme. Here, we discuss relevant examples, with emphasis on the rational engineering of anticoagulant thrombin mutants.

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Section: Allostery In Trypsin-like Enzymes: the E* And E Formsmentioning

confidence: 99%

Allostery in trypsin-like proteases suggests new therapeutic strategies

Gohara¹,

Di²

2011

Trends in Biotechnology

111

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“…Modeling was performed using MODELLER (63) with μIR fragments from PDB ID code 3W11 (11) and native insulin from PDB ID code 2G4M (64). Molecular dynamics calculations were performed with GROMACS (v4.5.5) (65) using the OPLS-aa force field (66,67).…”

Section: Methodsmentioning

confidence: 99%

Protective hinge in insulin opens to enable its receptor engagement

Menting

Yang

Chan

et al. 2014

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

149

257

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Insulin provides a classical model of a globular protein, yet how the hormone changes conformation to engage its receptor has long been enigmatic. Interest has focused on the C-terminal Bchain segment, critical for protective self-assembly in β cells and receptor binding at target tissues. Insight may be obtained from truncated "microreceptors" that reconstitute the primary hormone-binding site (α-subunit domains L1 and αCT). We demonstrate that, on microreceptor binding, this segment undergoes concerted hinge-like rotation at its B20-B23 β-turn, coupling reorientation of Phe B24 to a 60°rotation of the B25-B28 β-strand away from the hormone core to lie antiparallel to the receptor's L1-β 2 sheet. Opening of this hinge enables conserved nonpolar side chains (Ile A2 , Val A3 , Val B12 , Phe B24 , and Phe B25 ) to engage the receptor. Restraining the hinge by nonstandard mutagenesis preserves native folding but blocks receptor binding, whereas its engineered opening maintains activity at the price of protein instability and nonnative aggregation. Our findings rationalize properties of clinical mutations in the insulin family and provide a previously unidentified foundation for designing therapeutic analogs. We envisage that a switch between free and receptorbound conformations of insulin evolved as a solution to conflicting structural determinants of biosynthesis and function.diabetes mellitus | signal transduction | receptor tyrosine kinase | metabolism | protein structure H ow insulin engages the insulin receptor has inspired speculation ever since the structure of the free hormone was determined by Hodgkin and colleagues in 1969 (1, 2). Over the ensuing decades, anomalies encountered in studies of analogs have suggested that the hormone undergoes a conformational change on receptor binding: in particular, that the C-terminal β-strand of the B chain (residues B24-B30) releases from the helical core to expose otherwise-buried nonpolar surfaces (the detachment model) (3-6). Interest in the B-chain β-strand was further motivated by the discovery of clinical mutations within it associated with diabetes mellitus (DM) (7). Analysis of residuespecific photo-cross-linking provided evidence that both the detached strand and underlying nonpolar surfaces engage the receptor (8).The relevant structural biology is as follows. The insulin receptor is a disulfide-linked (αβ) 2 receptor tyrosine kinase (Fig. 1A), the extracellular α-subunits together binding a single insulin molecule with high affinity (9). Involvement of the two α-subunits is asymmetric: the primary insulin-binding site (site 1*) comprises the central β-sheet (L1-β 2 ) of the first leucine-rich repeat domain (L1) of one α-subunit and the partially helical Cterminal segment (αCT) of the other α-subunit (Fig. 1A) (10). Such binding initiates conformational changes leading to transphosphorylation of the β-subunits' intracellular tyrosine kinase (TK) domains. Structures of wild-type (WT) insulin (or analogs) bound to extracellular receptor fragments were recently...

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“…Thus, in conclusion, it can be made mandatory while studying any macromolecule that the structural determination is complemented with a long-wavelength data set, in addition to the soft X-ray molecular crystallography. The lectin contained two protein S atoms, one manganese ion, one calcium ion, one sodium ion and three partially occupied chloride ions (q = 0.85-0.65) 28 . 29 .…”

Section: Primary Structures Of Concanavalin A-like Lectins From Seedsmentioning

confidence: 99%

Untitled

2016

IJPSR

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Man has been dependent on plants for the treatment and cure of different diseases since time immemorial. Plants have served not only as a source of food and shelter but also as a very important source of different "portions" used for the treatment and cure of diseases. These portions are herbal or plant based mixtures made by traditional healers; who are individuals having a knowledge of medicinal plants of that particular area. In India, there are several thousands of medicinal plants, known to be used for specific diseases. Canavalia virosa is one such plant, known to be used for different curative purposes. However, it is more popular as a food plant as it is a tribal pulse. Canavalia virosa is a flowering plant belonging to the family Fabaceae. It is one of the lesser know vines from the genus Canavalia. It is not only important as an alternative protein source but also a promising medicinal plant as indicated by traditional medicine systems. This paper brings together some of the work carried out on this wonder plant by different researches.

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On the routine use of soft X-rays in macromolecular crystallography. Part IV. Efficient determination of anomalous substructures in biomacromolecules using longer X-ray wavelengths

Cited by 90 publications

References 33 publications

Allostery in trypsin-like proteases suggests new therapeutic strategies

Allostery in trypsin-like proteases suggests new therapeutic strategies

Protective hinge in insulin opens to enable its receptor engagement

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