RESUMEN
The human Gadd45 protein family plays critical roles in DNA repair, negative growth control, genomic stability, cell cycle checkpoints and apoptosis. Here we report the crystal structure of human Gadd45γ [corrected], revealing a unique dimer formed via a bundle of four parallel helices, involving the most conserved residues among the Gadd45 isoforms. Mutational analysis of human Gadd45γ [corrected] identified a conserved, highly acidic patch in the central region of the dimer for interaction with the proliferating cell nuclear antigen (PCNA), p21 and cdc2, suggesting that the parallel dimer is the active form for the interaction. Cellular assays indicate that: (1) dimerization of Gadd45γ [corrected] is necessary for apoptosis as well as growth inhibition, and that cell growth inhibition is caused by both cell cycle arrest and apoptosis; (2) a conserved and highly acidic patch on the dimer surface, including the important residues Glu87 and Asp89, is a putative interface for binding proteins related to the cell cycle, DNA repair and apoptosis. These results reveal the mechanism of self-association by Gadd45 proteins and the importance of this self-association for their biological function.
Asunto(s)
Péptidos y Proteínas de Señalización Intracelular/química , Secuencias de Aminoácidos , Animales , Apoptosis/efectos de la radiación , Proteína Quinasa CDC2 , Ciclo Celular , Supervivencia Celular , Cristalografía por Rayos X , Ciclina B/metabolismo , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Quinasas Ciclina-Dependientes , Células HeLa , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Ratones , Mutagénesis Sitio-Dirigida , Mutación Missense , Antígeno Nuclear de Célula en Proliferación/metabolismo , Unión Proteica , Dominios y Motivos de Interacción de Proteínas , Multimerización de Proteína , Estructura Cuaternaria de Proteína , Rayos UltravioletaRESUMEN
The relations of the binding free energies in a dataset of 69 protein complexes with the numbers of interfacial atom pairs, as well as with the atomic distances of the pairs, are analyzed. It is found that the interfacial main-chain atom pairs contribute more to the correlation than the interfacial side chain atom pairs do, and the polar atom pairs contribute more than the non-polar atom pairs do. Interfacial atom pairs with atomic distance in the range of 6-12 A are the most important to explain the differences in binding free energies in the datasets.
Asunto(s)
Complejos Multiproteicos/química , Interpretación Estadística de Datos , Unión ProteicaRESUMEN
2-Substituted 3-nitropropanoic acids were designed and synthesized as inhibitors against carboxypeptidase A (CPA). (R)-2-Benzyl- 3-nitropropanoic acid showed a potent inhibition against CPA (K(i)=0.15 microM). X-ray crystallography discloses that the nitro group well mimics the transition state occurred in the hydrolysis catalyzed by CPA, that is, an O,O'-bidentate coordination to the zinc ion and the two respective hydrogen bonds with Glu-270 and Arg-127. Because the nitro group is a planar species, we proposed (R)-2-benzyl-3-nitropropanoic acid as a pseudo-transition-state analog inhibitor against CPA.
Asunto(s)
Carboxipeptidasas A/antagonistas & inhibidores , Carboxipeptidasas A/metabolismo , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Nitrógeno/química , Zinc/química , Catálisis , Cristalografía por Rayos X , Inhibidores Enzimáticos/química , Modelos Moleculares , Estructura Molecular , Relación Estructura-ActividadRESUMEN
The number of atom-atom contacts in long distance can fit to the experimental binding energies in a dataset containing 151 experimental data with the correlation coefficient about 0.68. Based on this factor, a set of distance-dependent empirical potentials for various types of short-distance (2.4 A-5 A) contacts was obtained by guided fitting, i.e. a set of two parameters fitting. Incorporation of these short-distance potentials improved the correlation coefficients to 0.881.
Asunto(s)
Conformación Proteica , Proteínas , Bases de Datos de Proteínas , Modelos Teóricos , Datos de Secuencia Molecular , Unión Proteica , Proteínas/química , Proteínas/metabolismo , Estadística como Asunto , TermodinámicaRESUMEN
How protein molecules pack into a crystal remains problematic. Packing units are direct materials for packing into crystals. The group generator method is introduced for automatically identifying the packing unit. By introducing deviations into the nucleation stage of crystallization, we proved that these deviations diminish in further packing. This process illustrates how translation symmetries are achieved by the growing of nuclei. Two effects, the size effect and the close up effect, are found to behave differently in this process.
Asunto(s)
Núcleo Celular/metabolismo , Biosíntesis de Proteínas , Proteínas/metabolismo , Cristalización , Modelos Biológicos , Proteínas/química , Proteínas/genéticaRESUMEN
When amino acid residues are represented by parameters describing their side chain lengths and polarities, a sequence function defined as the sum of the first two sequence autocorrelation functions is found to be negatively and linearly correlated with the logarithms of folding rates of beta-proteins. The new function reveals new features in beta-protein folding: larger residues slow down the folding while alternative distribution of polar-non-polar residues accelerates the folding.
Asunto(s)
Conformación Proteica , Pliegue de Proteína , Proteínas/química , Secuencia de Aminoácidos , Matemática , Estructura Secundaria de ProteínaRESUMEN
Thioredoxin, DsbA, the N-terminal active-site domain a and the non-active-site domain b of protein-disulfide isomerase are all monomeric with a thioredoxin fold, and each exhibits low or no isomerase and chaperone activity. We have linked the N terminus of the above four monomers, individually, to the C terminus of the N-terminal domain of DsbC via the flexible linker helix of the latter to produce four domain hybrids, DsbCn-Trx, DsbCn-DsbA, DsbCn-PDIa, and DsbCn-PDIb. These four hybrid proteins form homodimers, and except for DsbCn-PDIb they exhibit new or greatly elevated isomerase as well as chaperone activity. Three-dimensional structure prediction indicates that all the four domain hybrids adopt DsbC-like V-shaped structure with a broad uncharged cleft between the two arms for binding of non-native protein folding intermediates. The results provide strong evidence that dimerization creates chaperone and isomerase activity for monomeric thiol-protein oxidases or reductases, and suggesting a pathway for proteins to acquire new functions and/or higher biological efficiency during evolution.
Asunto(s)
Chaperonas Moleculares/química , Sitios de Unión , Dicroismo Circular , Dimerización , Electroforesis en Gel de Poliacrilamida , Escherichia coli/metabolismo , Guanidina/química , Modelos Moleculares , Plásmidos/metabolismo , Unión Proteica , Conformación Proteica , Proteína Disulfuro Isomerasas/química , Pliegue de Proteína , Estructura Terciaria de Proteína , Tiorredoxinas/química , Factores de TiempoRESUMEN
It is found that the helix parameter (HP), which favors clustering of non-polar residues, is linearly correlated with the logarithms of rate constants of folding of small two-state alpha-helical proteins. The definition is HP = N(H)(-1) sigma [f(i)+ (f(i-1)+f(i+1))/2], where f(i)=1 or -1, if the i'th residue is hydrophobic or hydrophilic, respectively, N(H) is the number of hydrophobic residues and the summation is taken over the hydrophobic residues.
Asunto(s)
Conformación Proteica , Pliegue de Proteína , Proteínas/química , Algoritmos , Aminoácidos/química , Animales , Humanos , Modelos QuímicosRESUMEN
The crystal structure of pokeweed antiviral protein from seeds of Phytolacca americana (PAP-S) was solved at 1.8A. PAP-S is a one-chain ribosome-inactivating protein (RIP) and distinctively contains three well-defined N-acetylglucosamines, each covalently linked to an asparagine residue at positions, 10, 44, and 255, respectively. The high-resolution structure clearly shows the three mono-sugars to have either an alpha- or a beta-conformation. Two of sugars are located on the same side of the molecule with the active pocket. Except one hydrogen bond, there are no intermolecular interactions between the polypeptide chain and the sugars. Instead the sugar conformations appear to be stabilized by intermolecular interactions. The sugar structure defined at high resolution provides a structural basis for understanding their possible biological activity. The structural comparisons of PAP-S with other PAPs reveal that the major disparity of these homologous molecules is the different charge distribution on the upper right side of the front side near the active pocket. Based on the available structure of the 50S ribosomal subunit, the possible interactions between PAPs and the ribosome are discussed.