Search

The structure of BCL-w with a bound BID-BH3 peptide is determined using NMR spectroscopy and molecular docking. The interactions of BCL-w and the BID-BH3 peptide complex with dodecylphosphocholine micelles are characterized and demonstrated that the conformational change of BCL-w upon lipid binding occurs at the same time as the release and unfolding of the BH3 peptide.

Bacteriocins produced by latic acid bacteria are potent antimicrobial compounds that are active against closely related bacteria. The NMR solution structure of the 111-amino acid immunity protein for carnobacteriocin B2 (ImB2) is reported.

Changes in conformation in the PBX homeodomain and C-terminal extension after binding DNA and HOX-derived YPWM peptides are discussed. The structure of the free PBX homeodomain was found using NMR spectroscopy. PBX is in the PBC family of proteins with its homologues Drosophilia extradenticle and Caenorhabditis elegans ceh-20. They are in the three amino acid loop extension superclass of homeodomains.

The nuclear magnetic resonance solution structure of type IIa immunity protein, PisI, a 98-amino acid protein which is a group B immunity protein conferring immunity against piscicolin 126 (PisA) is reported. PisI, similar to ImB2, EntA-im, and PedB is observed to fold into a globular protein in aqueous solution and contains an antiparallel four-helix bundle.

The BCL-[x.sub.L] dimers are prepared by heating concentrated protein over 50 degree Celsius and the heat-induced mechanism of dimerization is done through [alpha]-helix swapping. The results have shown that the temperature can also induce three-dimensional domain swapping in BCL-2 family proteins

Multidimensional NMR studies on subtilosin A, a bacteriocin from Bacillus subtilis, are carried out to determine the complete primary and three-dimensional solution structure of peptide. Results depicted a cyclized peptide backbone with three cross-links formed between the sulfurs of Cys13, Cys 7, and Cys4 and the alpha-positions of Phe22, Thr28 and Phe31 that are showing L form of stereochemistry.

A new method to reduce dehydro side chains and reductively desulfurize lanthionine bridges found in lantibiotics, using nickel boride in the presence of deuterium gas, is reported. The application f a deuterating variant of the process, using NaBD (sub 4) in deuterated solvent, to lantibiotics leads to lanthionne desulfurization with the concomitant incorporation of a single deuterium atom at each of the residues participating in the sulfur bridge.

The dimerization of anti-apoptotic BCL-XL by three-dimensional domain swapping has recently been discovered at alkaline pH; however, the high energetic barrier between the dimer and monomer forms of BCL-XL prevents them from interconverting at room temperature and neutral pH. Here, we demonstrate that BCL-XL dimers can be easily prepared by heating concentrated protein above 50 °C. The 38 κDa BCL-XL dimer was fully characterized by multi-resonance nuclear magnetic resonance (NMR) spectroscopy, and the mechanism of dimerization by α-helix swapping was confirmed. Dimerization strongly affects the NMR signals from the turn between helices α5 and α6 of BCL-XL and a portion of the long loop between helices α1 and α2. Measurements of residual dipolar couplings demonstrate that the solution structure of the BCL-XL dimer is very close to the crystal structure. Dimer formation does not prevent tight binding of ligands to the hydrophobic cleft of BCL-xL; however, binding of a BID BH3—peptide or a polyphenol drug, gossypol, to BCL-XL significantly slowed monomer—dimer interconversion and is an example of the control of BCL protein oligomerization by ligand binding. [ABSTRACT FROM AUTHOR]

Bacteriocins produced by lactic acid bacteria are potent antimicrobial compounds which are active against closely related bacteria. Producer strains are protected against the effects of their cognate bacteriocins by immunity proteins that are located on the same genetic locus and are coexpressed with the gene encoding the bacteriocin. Several structures are available for class IIa bacteriocins; however, to date, no structures are available for the corresponding immunity proteins. We report here the NMR solution structure of the 111-amino acid immunity protein for carnobacteriocin B2 (ImB2). ImB2 folds into a globular domain in aqueous solution which contains an antiparallel four-helix bundle. Extensive packing by hydrophobic side chains in adjacent helices forms the core of the protein. The C-terminus, containing a fifth helix and an extended strand, is held against the four-helix bundle by hydrophobic interactions with helices 3 and 4. Most of the charged and polar residues in the protein face the solvent. Helix 3 is well-defined to residue 55, and a stretch of nascent helix followed by an unstructured loop joins it to helix 4. No interaction is observed between ImB2 and either carnobacteriocin B2 (CbnB2) or its precursor. Protection from the action of CbnB2 is only observed when ImB2 is expressed within the cell. The loop between helices 3 and 4, and a hydrophobic pocket which it partially masks, may be important for interaction with membrane receptors responsible for sensitivity to class IIa bacteriocins. [ABSTRACT FROM AUTHOR]

The complete primary and three-dimensional solution structures of subtilosin A (1), a bacteriocin from Bacillus subtilis, were determined by multidimensional NMR studies on peptide produced using isotopically labeled [(13)C,(15)N]medium derived from Anabaena sp. grown on sodium [(13)C]bicarbonate and [(15)N]nitrate. Additional samples of 1 were also generated by separate incorporations of [U-(13)C,(15)N]-L-phenylalanine and [U-(13)C,(15)N]-L-threonine using otherwise unlabeled media. The results demonstrate that in addition to having a cyclized peptide backbone (amide between N and C termini), three cross-links are formed between the sulfurs of Cys13, Cys7, and Cys4 and the alpha-positions of Phe22, Thr28, and Phe31, respectively. The stereochemistry of all residues in 1 except for the three modified ones was confirmed to be L by complete desulfurization with nickel boride, acid hydrolysis to the constituent amino acids, and conversion of these to the corresponding pentafluoropropanamide isopropyl esters for chiral GC MS analysis. The stereochemistry at the modified residues was determined by subjecting each of the eight possible stereoisomers of 1 to eight rounds of ARIA structure calculations, starting with the same NMR peak files and assignments. The stereoisomer with the l stereochemistry at Phe22 (alpha-R) and d stereochemistry at Thr28 (alpha-S) and Phe31 (alpha-S) (LDD isomer) fit the NMR data, giving the lowest energy family of structures with the best rmsd. Thus, biochemical formation of the unusual thio links proceeds with net retention of configuration at Phe22, and inversion at Thr28 and Phe31. Model amino acid derivatives bearing a sulfide moiety at the alpha-carbon were synthesized by reaction of the corresponding alpha-alkoxy compounds with benzyl thiol and SnCl(4). Separation of their pure stereoisomers and desulfurization with nickel boride demonstrated that the reduction of such compounds proceeds with epimerization, in contrast to the previously reported retention of stereochemistry for analogous reaction of steroidal sulfides. However, desulfurization of subtilosin A to cyclic peptide 14, which is inactive as an antimicrobial agent, occurs with inversion of stereochemistry at the alpha-carbons of Phe22 and Thr28 and with 4:1 retention at Phe31. This indicates that the desulfurization reaction proceeds via an N-acyl imine and that the structure of the surrounding peptide controls the geometry of reduction. Posttranslational linkage of a thiol to the alpha-carbon of an amino acid residue is unprecedented in ribosomally synthesized peptides or proteins, and very rare in secondary metabolites. Subtilosin A (1) represents a new class of bacteriocins.