Item 1: November 2005 Publications by Members:

1:  Koike C, Moore R, Negishi M.
 Localization of the nuclear receptor CAR at the cell membrane of mouse liver.
FEBS Lett. 2005 Nov 21; [Epub ahead of print] PMID: 16310787

2:  Huang CC, Tang M, Zhang MY, Majeed S, Montabana E, Stanfield RL, Dimitrov DS, Korber B, Sodroski J, Wilson IA, Wyatt R, Kwong PD.
 Structure of a V3-containing HIV-1 gp120 core.
Science. 2005 Nov 11;310(5750):1025-8. PMID: 16284180

3:  Guo H, Wlodawer A, Guo H.
 A general acid-base mechanism for the stabilization of a tetrahedral adduct in a serine-carboxyl peptidase: a computational study.
J Am Chem Soc. 2005 Nov 16;127(45):15662-3. PMID: 16277482

4:  Suh JY, Tang C, Cai M, Clore GM.
 Visualization of the phosphorylated active site loop of the cytoplasmic B
domain of the mannitol transporter II(Mannitol) of the Escherichia coli
phosphotransferase system by NMR spectroscopy and residual dipolar couplings.
J Mol Biol. 2005 Nov 11;353(5):1129-36. PMID: 16219324

5:  Louis JM, Bewley CA, Gustchina E, Aniana A, Clore GM.
 Characterization and HIV-1 fusion inhibitory properties of monoclonal Fabs obtained from a human non-immune phage library selected against diverse epitopes of the ectodomain of HIV-1 gp41.
J Mol Biol. 2005 Nov 11;353(5):945-51. PMID: 16216270

6:  Li M, Rasulova F, Melnikov EE, Rotanova TV, Gustchina A, Maurizi MR, Wlodawer A.
 Crystal structure of the N-terminal domain of E. coli Lon protease.
Protein Sci. 2005 Nov;14(11):2895-900. PMID: 16199667

7:  LaRonde-LeBlanc N, Wlodawer A.
 A family portrait of the RIO kinases.
J Biol Chem. 2005 Nov 11;280(45):37297-300. PMID: 16183636

8:  Burgess BR, Schuck P, Garboczi DN.
 Dissection of merozoite surface protein 3, a representative of a family of
Plasmodium falciparum surface proteins, reveals an oligomeric and highly
elongated molecule.
J Biol Chem. 2005 Nov 4;280(44):37236-45. PMID: 16135515

Item 2: Tips and Tricks

This section is always open for contributions. Click for Introduction and tips and tricks in Crystallization, Post-crystallization treatments for improving diffraction quality of protein crystals, Derivatization, Diffraction, Symmetry, Structure Solution, Structure Refinement, and Structure Analysis.

Item 3: Topic Discussion

Click for previous discussions on: PHASER, HKL2000, Parallel Protein Expression, Structural Genomics, NCS, Missing Atoms, Trends in Crystallography, and Absorption Correction.

PHASER: MR with Maximum Likelihood

Drs. Mariusz Jaskolski (UAM) & Alexander Wlodawer (NCI):

Although numerous crystal structures of several retroviral proteases (PR) have been solved in the past (e.g. from HIV, RSV, FIV, EIAV) and all these enzymes share significant sequence and structure homology, the new structure of human T-cell leukemia virus (HTLV) PR could not be solved (in our hands) by any of the standard MR programs (AMoRe, EPMR, MolRep). However, the structure was solved in a straightforward manner using the PHASER program, as implemented in ccp4i.

Some explanation of the failure of the "older" MR programs is provided by the comparatively low levels of sequence identity/similarity between the models and the target (ranging in simple CLUSTAL comparisons from 23.9/59.3% for FIV PR to 32.3/63.6% for HIV PR) and by the complicated architecture of the unknown structure (three homodimeric molecules forming a highly pseudosymmetric trimer), combined with high degree of uncertainty about the asymmetric unit contents.

In automatic runs, which included the full available resolution of 2.6 A, PHASER correctly solved the trimeric model, but failed (correctly again) to located any additional copies of the molecule due to packing considerations. The solutions could be unambiguously identified by their high Z-score parameter (> 10) for the more similar models (HIV PR, EIAV PR). The less similar models had Z-score values just below 8, and corresponded to a correct (7.96, RSV PR) or incorrect (7.80, FIV PR) solution. The quality of the solutions is also recognizable from the final LL-gain parameter, which at a value of 110 could distinguish between incorrect and correct solutions, increased to about 180 for "strong", unambiguous solutions (Z-score about 11), and rose sharply to 270 for the absolutely best case (Z-score 14.54).

In post factum analyses, the levels of sequence identity/similarity are reflected in the r.m.s. deviations between the Ca atoms of the corresponding models and the final HTLV PR structure, which are high even for the best models: HIV PR 1.6 A, EIAV PR 1.7 A, FIV PR 1.8 A, RSV PR 1.9 A.

In practical terms, the structure was cracked using the best available model of HIV PR determined at atomic resolution (Z-score 11.16). However, in a posteriori calculations the clearest solution was obtained with a medium-resolution HIV PR model, which in Ca comparison with the final target did not show any obvious superiority. This observation reinforces the notion that the performance of even the most robust MR algorithms may critically depend on the initial conditions and that, whenever available, different models should be tried.

Item 4: Lectures

Dr. Zbigniew Dauter's Lectures at the NIH (03/29-31)