Biomedical subjects

Maria Solà

Publications and source records attributed to Maria Solà.

4 recordsLinked to original sources

PubMed2006

The X-ray crystal structures of two constitutively active mutants of the Escherichia coli PhoB receiver domain give insights into activation.

The PhoR/PhoB two-component system is a key regulatory protein network enabling Escherichia coli to respond to inorganic phosphate (Pi) starvation conditions by turning on Pho regulon genes for more efficient Pi uptake and use of alternative phosphorus sources. Under environmental Pi depletion, the response regulator (RR) component, PhoB, is phosphorylated at the receiver domain (RD), a process that requires Mg(2+) bound at the active site. Phosphorylation of the RD relieves the inhibition of the PhoB effector domain (ED), a DNA-binding region that binds to Pho regulon promoters to activate transcription. The molecular details of the activation are proposed to involve dimerization of the RD and a conformational change in the RD detected by the ED. The structure of the PhoB RD shows a symmetrical interaction involving alpha1, loop beta5alpha5 and N terminus of alpha5 elements, also seen in the complex of PhoB RD with Mg(2+), in which helix alpha4 highly increases its flexibility. PhoB RD in complex with Mg(2+) and BeF(3) (an emulator of the phosphate moiety) undergoes a dramatic conformational change on helix alpha4 and shows another interaction involving alpha4, beta5 and alpha5 segments. We have selected a series of constitutively active PhoB mutants (PhoB(CA)) that are able to turn on the Pho regulon promoters in the absence phosphorylation and, as they cannot be inactivated, should therefore mimic the active RD state of PhoB and its functional oligomerisation. We have analysed the PhoB(CA) RD crystal structures of two such mutants, Asp53Ala/Tyr102Cys and Asp10Ala/Asp53Glu. Interestingly, both mutants reproduce the homodimeric arrangement through the symmetric interface encountered in the unbound and magnesium-bound wild-type PhoB RD structures. Besides, the mutant RD structures show a modified active site organization as well as changes at helix alpha4 that correlate with repositioning of surrounding residues, like the active-site events indicator Trp54, putatively redifining the interaction with the ED in the full-length protein.

Amino Acid Motifs↗

PubMed2006

Substrate specificity of a metalloprotease of the pappalysin family revealed by an inhibitor and a product complex.

Human pappalysin-1 is a multi-domain metalloprotease engaged in the homeostasis of insulin-like growth factors and the founding member of the pappalysin family within the metzincin clan of metalloproteases. We have recently identified an archaeal relative, ulilysin, encompassing only the protease domain. It is a 262-residue active protease with a novel 3D structure with two subdomains separated by an active-site cleft. Despite negligible overall sequence similarity, noticeable similarity is found with other metzincin prototypes, adamalysins/ADAMs and matrix metalloproteinases. Ulilysin has been crystallised in a product complex with an arginine-valine dipeptide occupying the active-site S(1') and S(2') positions and in a complex with the broad-spectrum hydroxamic acid-based metalloprotease inhibitor, batimastat. This molecule inhibits mature ulilysin with an IC(50) value of 61 microM under the conditions assayed. The binding of batimastat to ulilysin evokes binding to vertebrate matrix metalloproteases but is much weaker. These data give insight into substrate specificity and mechanism of action and inhibition of the novel pappalysin family.

Amino Acid Sequence↗

PubMed2006

The cofactor-induced pre-active conformation in PhoB.

PhoB is an Escherichia coli transcription factor from a two-component signal transduction system that is sensitive to limiting environmental phosphate conditions. It consists of an N-terminal receiver domain (RD) and a C-terminal DNA-binding domain. The protein is activated upon phosphorylation at the RD, an event that depends on Mg(2+) binding. The structure of PhoB RD in complex with Mg(2+) is presented, which shows three protomers in the asymmetric unit that interact across two different surfaces. One association is symmetric and has been described as a non-active dimerization contact; the other involves the alpha4-beta5-alpha5 interface and recalls the contact found in activated PhoB. However, here this last interaction is not perfectly symmetric and helix alpha4, which in the activated molecule undergoes a helical shift, becomes strongly destabilized in one of the interacting monomers. All protomers bind the cation in a similar manner but, interestingly, at the prospective binding site for the phosphate moiety the side chains of either Glu88 (in helix alpha4) or Trp54 alternate and interact with active-site atoms. When Glu88 is inside the cavity, helix alpha4 is arranged similarly to the unliganded wild-type structure. However, if Trp54 is present, the helix loses its contacts with the active-site cavity and vanishes. Accordingly, the presence of Trp54 in the active site induces a flexible state in helix alpha4, potentially allowing a helical shift that phosphorylation would eventually stabilize.

Bacterial Proteins↗

PubMedDate not supplied

Papaya glutamine cyclotransferase shows a singular five-fold beta-propeller architecture that suggests a novel reaction mechanism.

Cyclisation of N-terminal glutamine and/or glutamate to yield pyroglutamate is an essential posttranslational event affecting a plethora of bioactive peptides and proteins. It is directly linked with pathologies ranging from neurodegenerative diseases to inflammation and several types of cancers. The reaction is catalysed by ubiquitous glutaminyl cyclotransferases (QCs), which present two distinct prototypes. Mammalian QCs are zinc-dependent enzymes with an alpha/beta-hydrolase fold. Here we present the 1.6-A-resolution structure of the other prototype, the plant analogue from Carica papaya (PQC). The hatbox-shaped molecule consists of an unusual five-fold beta-propeller traversed by a central channel, a topology that has hitherto been described only for some sugar-binding proteins and an extracellular nucleotidase. The high resistance of the enzyme to denaturation and proteolytic degradation is explained by its architecture, which is uniquely stabilised by a series of tethering elements that confer rigidity. Strikingly, the N-terminus of PQC specifically interacts with residues around the entrance to the central channel of a symmetry-related molecule, suggesting that this location is the putative active site. Cyclisation would follow a novel general-acid/base working mechanism, pivoting around a strictly conserved glutamate. This study provides a lead structure not only for plant QC orthologues, but also for bacteria, including potential human pathogens causing diphtheria, plague and malaria.

Amino Acid Sequence↗