Journal of Bioinformatics and Computational Biology, December 2009, p. 1031-1037, Vol. 7, No. 6

Exploring the Protein Landscape in Ramachandran Space: It's not just Psi-Phi.

William Krivan and Darrick Carter

Protein Advances Inc., Seattle, Washington Received 23 April 2009/ Revised 11 August 2009/ Accepted 11 August 2009

Most methods for the structural comparison of proteins utilize molecular coordinates in the three-dimensional physical space. Recently, a group has presented an elegant novel approach based on the characterization of protein shape in terms of backbone torsion angles. They have demonstrated considerable success in direct comparisons with other techniques, and their method lends itself to rapid screening of structural information from rapidly growing databases. We think that the torsion angle approach can be further strengthened by refining the distance notion that forms the basis of the computational scheme. In particular, we are suggesting to compute the distance along the path that minimizes the transition cost between aligned pairs of angles and therefore likely provides a more meaningful representation of distances between points in Ramachandran space.

Infection and Immunity, September 2008, p. 3967-3974, Vol. 76, No. 9

Immunological Dominance of Trypanosoma cruzi Tandem Repeat Proteins

Yasuyuki Goto,^1* Darrick Carter,^1,2 and Steven G. Reed¹

Infectious Disease Research Institute,¹ Protein Advances Inc., Seattle, Washington² Received 16 May 2008/ Returned for modification 20 June 2008/ Accepted 30 June 2008

Proteins with tandem repeat (TR) domains have been found invarious protozoan parasites, often acting as targets of B-cellresponses. However, the extent of the repeats within Trypanosomacruzi, the causative agent of Chagas? disease, has notbeen examined well. Here, we present a systematic survey ofthe TR genes found in T. cruzi, in comparison with other organisms.Although the characteristics of TR genes varied from organismto organism, the presence of genes having large TR domains wasunique to the trypanosomatids examined, including T. cruzi.Sequence analyses of T. cruzi TR genes revealed their divergency;they do not share such characteristics as sequence similarityor biased cellular location predicted by the presence of a signalsequence or transmembrane domain(s). In contrast, T. cruzi TRproteins seemed to possess significant antigenicity. A numberof previously characterized T. cruzi antigens were detectedby this computational screening, and several of those antigenscontained a large TR domain. Further analyses of the T. cruzigenome demonstrated that previously uncharacterized TR proteinsin this organism may also be immunodominant. Taken together,T. cruzi is rich in large TR domain-containing proteins with immunological significance; it is worthwhile further analyzingsuch characteristics of TR proteins as the copy number and consensussequence of the repeats to determine whether they might contributeto the biological variability of T. cruzi strains with regardto induced immunological responses, host susceptibility, diseaseoutcomes, and pathogenicity.

Clin Vaccine Immunol. 2008 Aug 20. [Epub ahead of print]

Selection of antigens and prototype test development for a point-of-care leprosy diagnosis.

Duthie MS, Ireton GC, Kanaujia GV, Goto W, Liang H, Bhatia A, Busceti JM, Macdonald M, Neupane KD, Ranjit C, Sapkota BR, Balagon M, Esfandiari J, Carter D, Reed SG.

Infectious Disease Research Institute, 1124 Columbia St, Suite 400, Seattle, WA 98104; ChemBio Diagnostic Systems Inc., 3661 Horseblock Road, Medford, NY 11763; Mycobacterial Research Laboratory, Anandaban Hospital, Kathmandu, Nepal; Leonard Wood Memorial Center for Leprosy Research, Cebu City, Philippines; and Protein AI, 1124 Columbia St, Seattle, WA 98104.

Leprosy can be a devastating chronic infection that causes nerve function impairment and associated disfigurement. Despite the recent reduction in the number of registered worldwide leprosy cases as a result of the widespread use of multi-drug therapy, the number of new cases detected each year remains relatively stable. The diagnosis of leprosy is currently based on the appearance of clinical signs and requires expert clinical, as well as labor intensive and time consuming laboratory or histological evaluation. For the purpose of developing an effective, simple, rapid, and low cost diagnostic alternative, we have analyzed the serologic antibody response to identify M. leprae proteins that are recognized by leprosy patients. More than 100 recombinant antigens were analyzed in protein array format to select those with discriminatory properties for leprosy diagnosis. As expected, multibacillary leprosy patients recognized more antigens with stronger antibody responses than paucibacillary leprosy patients. Our data indicate, however, that multibacillary patients can be distinguished from paucibacillary patients, and both of these groups can be segregated from endemic control groups. We went on to confirm the diagnostic properties of antigens ML0405, ML2331, and the LID-1 fusion construct of these two proteins by ELISA. We then demonstrated the performance of these antigens in rapid test formats with a goal of developing a point-of-care diagnostic test. A serological diagnostic test capable of identifying and allowing treatment of leprosy could reduce transmission, prevent functional disabilities and stigmatizing deformities, and facilitate leprosy eradication.

Abstract presented at the 43rd Annual Tuberculosis and Leprosy Conference, July 10^th 2008

In silicogenome reduction, a method for synthesizing the leprosy proteome?

By Darrick Carter^1,2, Steven G. Reed¹, Malcolm Duthie^1,2, and William Krivan²

From the ¹Infectious Disease Research Institute, 1124 Columbia St, Suite 400, Seattle, WA 98104, ²Protein Advances Inc. (ProteinAI), 1124 Columbia St, Suite 401, Seattle, WA 98104.

The genome of M. leprae has been sequenced in its entirety and exhibits partial or total loss of multiple biosynthetic pathways. This decay of the genome is reflected by the low number of remaining expressed coding sequences and the array of residual pseudogenes. Specifically, the 3.27 megabases that make up the genetic information of the bacterium, contain 1,604 predicted coding sequences and 1,116 pseudogenes. Using software written at ProteinAI, we have broken the proteome into 36,236 overlapping 20-mer peptides. Subsequently, we constructed a symmetric matrix representing the pairwise Hamming distance between the peptides. The Hamming distance is a concept from information theory, and was used as a measure for similarity in order to identify pools of related peptides. Finding the minimal number of pools for a chosen distance threshold is an optimization problem known to be NP-hard, therefore we cannot expect to be able to find an efficient algorithm to solve the problem exactly. For this reason, an approximation method, the ?Greedy? algorithm1, - guaranteed to find a reduced set number with maximum 2 fold error - had to be used. This allowed for the grouping of related peptides into 334 pools with a maximum distance of 16. Practically, this could allow for complete leprosy proteome peptide screening in two steps each using a single microtiter plate. The same approach is now being extended to tuberculosis and other organisms.


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Science Literature Google Scholar PubMed at NIH	Publications from Protein AI Journal of Bioinformatics and Computational Biology, December 2009, p. 1031-1037, Vol. 7, No. 6 Exploring the Protein Landscape in Ramachandran Space: It's not just Psi-Phi. William Krivan and Darrick Carter Protein Advances Inc., Seattle, Washington Received 23 April 2009/ Revised 11 August 2009/ Accepted 11 August 2009 Most methods for the structural comparison of proteins utilize molecular coordinates in the three-dimensional physical space. Recently, a group has presented an elegant novel approach based on the characterization of protein shape in terms of backbone torsion angles. They have demonstrated considerable success in direct comparisons with other techniques, and their method lends itself to rapid screening of structural information from rapidly growing databases. We think that the torsion angle approach can be further strengthened by refining the distance notion that forms the basis of the computational scheme. In particular, we are suggesting to compute the distance along the path that minimizes the transition cost between aligned pairs of angles and therefore likely provides a more meaningful representation of distances between points in Ramachandran space. Infection and Immunity, September 2008, p. 3967-3974, Vol. 76, No. 9 Immunological Dominance of Trypanosoma cruzi Tandem Repeat Proteins Yasuyuki Goto,^1* Darrick Carter,^1,2 and Steven G. Reed¹ Infectious Disease Research Institute,¹ Protein Advances Inc., Seattle, Washington² Received 16 May 2008/ Returned for modification 20 June 2008/ Accepted 30 June 2008 Proteins with tandem repeat (TR) domains have been found invarious protozoan parasites, often acting as targets of B-cellresponses. However, the extent of the repeats within Trypanosomacruzi, the causative agent of Chagas? disease, has notbeen examined well. Here, we present a systematic survey ofthe TR genes found in T. cruzi, in comparison with other organisms.Although the characteristics of TR genes varied from organismto organism, the presence of genes having large TR domains wasunique to the trypanosomatids examined, including T. cruzi.Sequence analyses of T. cruzi TR genes revealed their divergency;they do not share such characteristics as sequence similarityor biased cellular location predicted by the presence of a signalsequence or transmembrane domain(s). In contrast, T. cruzi TRproteins seemed to possess significant antigenicity. A numberof previously characterized T. cruzi antigens were detectedby this computational screening, and several of those antigenscontained a large TR domain. Further analyses of the T. cruzigenome demonstrated that previously uncharacterized TR proteinsin this organism may also be immunodominant. Taken together,T. cruzi is rich in large TR domain-containing proteins with immunological significance; it is worthwhile further analyzingsuch characteristics of TR proteins as the copy number and consensussequence of the repeats to determine whether they might contributeto the biological variability of T. cruzi strains with regardto induced immunological responses, host susceptibility, diseaseoutcomes, and pathogenicity. Clin Vaccine Immunol. 2008 Aug 20. [Epub ahead of print] Selection of antigens and prototype test development for a point-of-care leprosy diagnosis. Duthie MS, Ireton GC, Kanaujia GV, Goto W, Liang H, Bhatia A, Busceti JM, Macdonald M, Neupane KD, Ranjit C, Sapkota BR, Balagon M, Esfandiari J, Carter D, Reed SG. Infectious Disease Research Institute, 1124 Columbia St, Suite 400, Seattle, WA 98104; ChemBio Diagnostic Systems Inc., 3661 Horseblock Road, Medford, NY 11763; Mycobacterial Research Laboratory, Anandaban Hospital, Kathmandu, Nepal; Leonard Wood Memorial Center for Leprosy Research, Cebu City, Philippines; and Protein AI, 1124 Columbia St, Seattle, WA 98104. Leprosy can be a devastating chronic infection that causes nerve function impairment and associated disfigurement. Despite the recent reduction in the number of registered worldwide leprosy cases as a result of the widespread use of multi-drug therapy, the number of new cases detected each year remains relatively stable. The diagnosis of leprosy is currently based on the appearance of clinical signs and requires expert clinical, as well as labor intensive and time consuming laboratory or histological evaluation. For the purpose of developing an effective, simple, rapid, and low cost diagnostic alternative, we have analyzed the serologic antibody response to identify M. leprae proteins that are recognized by leprosy patients. More than 100 recombinant antigens were analyzed in protein array format to select those with discriminatory properties for leprosy diagnosis. As expected, multibacillary leprosy patients recognized more antigens with stronger antibody responses than paucibacillary leprosy patients. Our data indicate, however, that multibacillary patients can be distinguished from paucibacillary patients, and both of these groups can be segregated from endemic control groups. We went on to confirm the diagnostic properties of antigens ML0405, ML2331, and the LID-1 fusion construct of these two proteins by ELISA. We then demonstrated the performance of these antigens in rapid test formats with a goal of developing a point-of-care diagnostic test. A serological diagnostic test capable of identifying and allowing treatment of leprosy could reduce transmission, prevent functional disabilities and stigmatizing deformities, and facilitate leprosy eradication. Abstract presented at the 43rd Annual Tuberculosis and Leprosy Conference, July 10^th 2008 In silicogenome reduction, a method for synthesizing the leprosy proteome? By Darrick Carter^1,2, Steven G. Reed¹, Malcolm Duthie^1,2, and William Krivan² From the ¹Infectious Disease Research Institute, 1124 Columbia St, Suite 400, Seattle, WA 98104, ²Protein Advances Inc. (ProteinAI), 1124 Columbia St, Suite 401, Seattle, WA 98104. The genome of M. leprae has been sequenced in its entirety and exhibits partial or total loss of multiple biosynthetic pathways. This decay of the genome is reflected by the low number of remaining expressed coding sequences and the array of residual pseudogenes. Specifically, the 3.27 megabases that make up the genetic information of the bacterium, contain 1,604 predicted coding sequences and 1,116 pseudogenes. Using software written at ProteinAI, we have broken the proteome into 36,236 overlapping 20-mer peptides. Subsequently, we constructed a symmetric matrix representing the pairwise Hamming distance between the peptides. The Hamming distance is a concept from information theory, and was used as a measure for similarity in order to identify pools of related peptides. Finding the minimal number of pools for a chosen distance threshold is an optimization problem known to be NP-hard, therefore we cannot expect to be able to find an efficient algorithm to solve the problem exactly. For this reason, an approximation method, the ?Greedy? algorithm1, - guaranteed to find a reduced set number with maximum 2 fold error - had to be used. This allowed for the grouping of related peptides into 334 pools with a maximum distance of 16. Practically, this could allow for complete leprosy proteome peptide screening in two steps each using a single microtiter plate. The same approach is now being extended to tuberculosis and other organisms.

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