No abstract

p70 ribosomal S6 kinase (p70S6K) is a downstream effector of the mTOR signaling pathway involved in cell proliferation, cell growth, cell-cycle progression, and glucose homeostasis. Multiple phosphorylation events within the catalytic, autoinhibitory, and hydrophobic motif domains contribute to the regulation of p70S6K. We report the crystal structures of the kinase domain of p70S6K1 bound to staurosporine in both the unphosphorylated state and in the 3-phosphoinositide-dependent kinase-1-phosphorylated state in which Thr-252 of the activation loop is phosphorylated. Unphosphorylated p70S6K1 exists in two crystal forms, one in which the p70S6K1 kinase domain exists as a monomer and the other as a domain-swapped dimer. The crystal structure of the partially activated kinase domain that is phosphorylated within the activation loop reveals conformational ordering of the activation loop that is consistent with a role in activation. The structures offer insights into the structural basis of the 3-phosphoinositide-dependent kinase-1-induced activation of p70S6K and provide a platform for the rational structure-guided design of specific p70S6K inhibitors.The ribosomal S6 kinase family belongs to the AGC 3 subfamily of serine-threonine protein kinases. In humans two forms of p70 ribosomal S6 kinases (S6K1 and S6K2) have been reported that are encoded by two different genes (RPS6KB1 and RPS6KB2), respectively (1, 2). RPS6KB1 encodes two isoforms that differ only at the N termini by 23 amino acid residues (2). The longer form of S6K1 contains an N-terminal nuclear localization signal, whereas the shorter isoform of S6K1 predominantly localizes in the cytosol.Several substrates of p70S6K have been identified including 40 S ribosomal protein S6, insulin substrate (IRS1), preapoptotic protein BAD, eukaryotic initiation factor (elF4B), eukaryotic elongation factor (eEF2K) and cAMP-response element modulator (CREMt) (3). The most studied substrate is the 40 S ribosomal protein S6, a major component of the machinery involved in protein synthesis in mammalian cells, suggesting that p70S6K plays a role in regulating translation.Several observations suggest a role for p70S6K in cancer (4, 5). For example, upstream regulators of p70S6K are deregulated in multiple types of cancer, and gene and protein overexpression is observed in various cancers (4, 5). In addition, p70S6K is also a downstream kinase of insulin receptor-mediated signaling and is a potential therapeutic target for the management of obesity and diabetes as shown by enhanced metabolic rate and insulin sensitivity in p70S6K knock-out mice (4, 5).The activation of p70S6K requires multiple phosphorylation events in both the kinase and autoinhibitory domains (Fig. 1). The C-terminal autoinhibitory domain, which is believed to block phosphorylation within the hydrophobic motif and the activation loop, is phosphorylated by upstream kinases such as ERK (6, 7). Other activating phosphorylation events occur at Thr-412 in the hydrophobic motif by mTOR (mammalian target of rapam...

In the paper by Sunami et al. [(2004), Acta Cryst. D60, 90±96] an incorrect version of Table 2 was published. The correct version is given here. References

Recent advances in neutron crystallographic studies have provided structural bases for quantum behaviors of protons observed in enzymatic reactions. Thus, we resolved the neutron crystal structure of a bacterial copper (Cu) amine oxidase (CAO), which contains a prosthetic Cu ion and a protein-derived redox cofactor, topa quinone (TPQ). We solved hitherto unknown structures of the active site, including a keto/enolate equilibrium of the cofactor with a nonplanar quinone ring, unusual proton sharing between the cofactor and the catalytic base, and metal-induced deprotonation of a histidine residue that coordinates to the Cu. Our findings show a refined active-site structure that gives detailed information on the protonation state of dissociable groups, such as the quinone cofactor, which are critical for catalytic reactions.

A DNA fragment d(GCGAAAGCT), known to adopt a stable mini-hairpin structure in solution, has been crystallized in the space group I4(1)22 with the unit-cell dimensions a = b = 53.4 A and c = 54.0 A, and the crystal structure has been determined at 2.5 A resolution. The four nucleotide residues CGAA of the first half of the oligomer form a parallel duplex with another half through the homo base pairs, C2:C2+ (singly-protonated between the Watson- Crick sites), G3:G3 (between the minor groove sites), A4:A4 (between the major groove sites) and A5:A5 (between the Watson-Crick sites). The two strands remaining in the half of the parallel duplex are split away in different directions, and they pair in an anti-parallel B-form duplex with the second half extending from a neighboring parallel duplex, so that an infinite column is formed in a head-to-tail fashion along the c-axis. It seems that a hexa-ammine cobalt cation supports such a branched and bent conformation of the oligomer. One end of the parallel duplex is stacked on the corresponding end of the adjacent parallel duplex; between them, the guanine base of the first residue is stacked on the fourth ribose of another duplex.