Protein structure prediction using a combination of sequence-based alignment, constrained energy minimization, and structural alignment.

We present a novel approach to protein structure prediction in which fold recognition techniques are combined with ab initio folding methods. Based on the predicted secondary structure, one of two different protocols is followed. For mostly alpha proteins, global optimization and sampling of a statistical energy function is used to generate many low-energy structures; these structures are then screened against a fold library. Any structural matches are then selected for further refinement. For proteins predicted to have significant beta-content, sequence and secondary structure-based alignment is used to identify candidate templates; spatial constraints are then extracted from these templates and used, along with the statistical energy function, in the global sampling and optimization program. Successes and failures of both protocols are discussed.

Cloning and genomic organization of beclin 1, a candidate tumor suppressor gene on chromosome 17q21.

The beclin 1 (BECN1) gene encodes a 60-kDa coiled-coil protein that interacts with the prototypic apoptosis inhibitor Bcl-2. Previous studies indicate that beclin 1 maps to a region approximately 150 kb centromeric to BRCA1 on chromosome 17q21 that is commonly deleted in breast, ovarian, and prostate cancer. The complete cDNA sequence of beclin 1 encodes a 2098-bp transcript, with a 120-bp 5' UTR, 1353-bp coding region, and 625-bp 3' UTR. Hybridization screening of a human genomic PAC library identified PAC 452O8, which contains the complete beclin 1 gene. Determination of the exon-intron structure of beclin 1 reveals 12 exons, ranging from 61 to 794 bp, which extend over 12 kb of the human genome. FISH analysis of human breast carcinoma cell lines using PAC 452O8 as probe identified allelic beclin 1 deletions in 9 of 22 cell lines. Sequencing of genomic DNA from 10 of these cell lines revealed no mutations in coding regions or splice junctions. Additionally, Northern blot analysis of 11 cell lines did not identify any abnormalities in beclin 1 transcripts. These results indicate that human breast carcinoma cell lines frequently contain allelic deletions of beclin 1, but not beclin 1 coding mutations.