Clone-by-clone sequencing This approach means that the chromosomes were mapped and then split up into sections.

A rough map was drawn for each of these sections, and then the sections themselves were split into smaller bits, with plenty of overlap between each of the bits. Each of these smaller bits would be sequenced, and the overlapping bits would be used to put the genome jigsaw back together again. First, by mapping the genome, researchers produce, at an early stage, a genetic resource that can be used to map genes. In addition, because every DNA sequence is derived from a known region, it is relatively easy to keep track of the project and to determine where there are gaps in the sequence. Moreover, assembly of relatively short regions of DNA is an efficient step. However, mapping can be a time-consuming, and costly, process.

Whole genome shotgun sequencing The alternative to the clone-by-clone approach is the 'bottom-up' whole genome shotgun (WGS) sequencing. Shotgun sequencing was developed by Fred Sanger in 1982. First, all the DNA is first broken into fragments. The fragments are then sequenced at random and assembled together by looking for overlaps. In recent approaches, libraries have been made from DNAfragments of 2000 base pairs and of 10,000 base pairs in length. The two sizes of fragment provide complementary results; bysequencing the ends of the fragments, each provides information about DNA sequences separated by known distances. Computer analysis is used to search the sequences for overlaps. The advantage of the whole-genome shotgun is that it requires no prior mapping. Its disadvantage is that large genomes need vast amounts of computing power and sophisticated software to reassemble the genome from its fragments. To sequence the genome from a mammal (billions of bases long), you need about about 60,000,000 individual DNA sequence reads. Reassembling these sequenced fragments requires huge investments in IT, and, unlike the cloneby-clone approach, assemblies can't be produced until the end of the project. Whole genome shotgun for large genomes is especially valuable if there is an existing 'scaffold' of organized sequences, localized to the genome, derived from other projects. When the whole genome shotgun data are laid on the 'scaffold' sequence, it is easier to resolve ambiguities. Today, whole genome shotgun is used for most bacterial genomes and as a 'top-up' of sequence data for many other genome projects.

The era of OMICS & Post genomic ERA:

The era of omics approaches has produced a huge amount of data. For instance, in a microarray assay the differential expression of hundreds of genes is evaluated. The next generation sequencing makes possible to sequence entire genomes. Mass spectrometry and proteomic protocols produce data at protein level. The high-trough put screening allows analyzing thousands of compounds in few experiments. Even if the data, protocols and aim of the study are usually published, the majority of data is not used in the work that has produced them. Fortunately, in the supplementary figures these data are always available for other groups that may be interested in. The next future of biological science will move towards the validation of data from omics approaches. Indeed, the limited economical resources will make difficult for a laboratory to effort the expenses of big experiments with thousands of samples. It will be reasonable to start novel studies based on data already published. Each group is normally specialized and has specific competences that can be fundamental to validate and complete the results produced by others in omics protocols. This novel approach will favor collaboration between groups and will improve the usefulness of scientific data. Future of Bioinformatics: We are currently witnessing a technological revolution. With the increase of sequencing projects, bioinformatics continues to make considerable progress in biology by providing scientists with access to the genomic information. This progress is especially contributed by the Human Genome Project. The information obtained with the help of Bioinformatics tools furthers our understanding of various genetic and other diseases and helps identify new drug targets. With technological developments of the Internet, scientists are now able to freely access volumes of such biological information, which enables the advancement of scientific discoveries in biomedicine.

In spite of being young, the science of Bioinformatics exhibits tremendous potential for playing a major role in the future development of science and technology. This is evident from the fact that modern biology and related sciences are increasingly becoming dependent on this new technology. It is expected that Bioinformatics will especially contribute in the future as the leading edge in biomedicine to pharmaceutical companies by expediently yielding a greater quantity of lead drugs for therapy.

Molecular Modeling Experimentally resolved structures of proteins, RNA, and Database (MMDB) DNA, derived from the Protein Data Bank (PDB), with value-addedfeatures such as explicit chemical graphs, computationally identified 3D domains (compact substructures) that are used to identify similar 3D structures, as well as links to literature, similar sequences, information about chemicals bound to the structures, and more. These connections make it possible, for example, tofind 3D structures for homologs of a protein sequence of interest, then interactively view the sequence-structure relationships,active sites, bound chemicals, journal articles, and more.

Medline: MEDLINE, the primary component of PubMed, is the NLM's premier citation database. MEDLINE's subject scope is biomedicine and health, broadly defined to encompass those areas of the life sciences, behavioral sciences, chemical sciences, and bioengineering. MEDLINE citation data comprises approximately 91% of the data distributed to MEDLINE/PubMed licensees. The other categories of records that reside in PubMed and are distributed to licensees are: a. the out-of-scope citations from a few selectively indexed MEDLINE journals (primarily covering general science and general chemistry) for which the life sciences articles are indexed for MEDLINE; b. in-process citations which provide a record for an article before it is indexed with MeSH and added to MEDLINE or converted to out-of-scope status; c. some citations in the OLDMEDLINE subset that have not yet been updated with current vocabulary and converted to MEDLINE status;

d. some citations in PubMed that precede the date a journal was selected for MEDLINE indexing if the citations are submitted after late 2003 to NLM electronically by the publishers; e. starting in summer 2005, prospective citations to articles from non-MEDLINE journals that submit full text to PubMed Central and are thus cited in PubMed. NLM now exports over 98% of PubMed records to MEDLINE/PubMed licensees. The approximately 2% of the records not exported to MEDLINE/PubMed licensees are those tagged [PubMed - as supplied by publisher] in PubMed. These are mostly records recently added directly to PubMed via electronic submission from a publisher, most of which are soon to proceed to the next stage, at which time the record will be exported. Most other non-exported records are either: older citations for the relatively few non-MEDLINE journals in PubMed; citations to MEDLINE journals prior to their date of selection for MEDLINE which were submitted electronically by the publishers before late July 2003; or citations electronically submitted for articles that appear on the Web in advance of the journal issue's release (i.e., ahead of print citations). Following publication of the completed issue, the item will be queued for issue level review and released in In-Data-Review status. All "publisher-supplied" citations have not been reviewed for accurate bibliographic data. PUBMED: PubMed comprises more than 23 million citations for biomedical literature from MEDLINE, life science journals, and online books. Citations may include links to full-text content from PubMed Central and publisher web sites. PubMed comprises over 22 million citations for biomedical literature from MEDLINE, life science journals, and online books. PubMed citations and abstracts include the fields of biomedicine and health, covering portions of the life sciences, behavioral sciences, chemical sciences, and bioengineering. PubMed also provides access to additional relevant web sites and links to the other NCBI molecular biology resources. PubMed is a free resource that is developed and maintained by the National Center for Biotechnology Information (NCBI), at the U.S. National Library of Medicine (NLM), located at the National Institutes of Health (NIH). Publishers of journals can submit their citations to NCBI and then provide access to the full-text of articles at journal web sites using LinkOut. Phylogenetic Analysis A dendrogram is a broad term for the diagrammatic representation of a phylogenetic tree. phylogenetic tree, also called Dendrogram, a diagram showing the evolutionary interrelations of a group of organisms derived from a common ancestral form. The ancestor is in the tree trunk; organisms that have arisen from it are placed at the ends of tree branches. The distance of one group from the other groups indicates the degree of relationship; i.e., closely

related groups are located on branches close to one another. Phylogenetic trees, although speculative, provide a convenient method for studying phylogenetic relationships.

A cladogram is a phylogenetic tree formed using cladistic methods. This type of tree only represents a branching pattern; i.e., its branch spans do not represent time or relative amount of character change. Phylogenetic trees that have clearly implied patterns of ancestry and descent, and a relative time axis. A cladogram is one kind of phylogenetic tree, a common ancestry tree. For some phylogenetic systematists, specifically those that some have termed Transformed Cladistics, the cladogram is the basic unit of analysis and is held to be fundamentally different from a phylogenetic tree. Specifically, it is only a graphic summarizing derived characters shared taxa with no necessary connotation of common ancestry, process of descent (evolution), or relative time axis. (see also phylogenetic tree)

A phylogram is a phylogenetic tree that has branch spans proportional to the amount of character change.

A chronogram is a phylogenetic tree that explicitly represents evolutionary time through its branch spans. Rooted tree A rooted phylogenetic tree is a directed tree with a unique node corresponding to the (usually imputed) most recent common ancestor of all the entities at the leavesof the tree. The most common method for rooting trees is the use of an uncontroversial outgroupclose enough to allow inference from sequence or trait data, but far enough to be a clear outgroup. Unrooted tree Unrooted trees illustrate the relatedness of the leaf nodes without making assumptions about ancestry at all. While unrooted trees can always be generated from rooted ones by simply omitting the root, a root cannot be inferred from an unrooted tree without some means of identifying ancestry; this is normally done by including an outgroup in the input data or introducing additional assumptions about the relative rates of evolution on each branch, such as an application of themolecular clock hypothesis. Figure 2 depicts an unrooted phylogenetic tree formyosin, a superfamily of proteins.

molecular clock: a measure of evolutionary change over time at the molecular level that is based on the theory that specific DNA sequences or the proteins they encode spontaneously mutate at constant rates and that is used chiefly for estimating how long ago two related organisms diverged from a common ancestor. It is used to estimate the time of occurrence of events called speciation or radiation. It is sometimes called a gene clock or evolutionary clock. The molecular clock technique is an important tool in molecular systematics, the use of molecular genetics information to determine the correct scientific classification of organisms or to study variation in selective forces. Knowledge of approximately constant rate of molecular evolution in particular sets of lineages also facilitates establishing the dates of phylogenetic events, including those not documented by fossils, such as the divergence of living taxa and the formation of the phylogenetic tree. But in these cases especially over long stretches of time the limitations of MCH (above) must be considered; such estimates may be off by 50% or more.

Genome annotation: The molecular clock technique is an important tool in molecular systematics, the use of molecular genetics information to determine the correct scientific classification of organisms or to study variation in selective forces. Knowledge of approximately constant rate of molecular evolution in particular sets of lineages also facilitates establishing the dates of phylogenetic events, including those not documented by fossils, such as the divergence of living taxa and the formation of the phylogenetic tree. But in these cases especially over long stretches of time the limitations of MCH (above) must be considered; such estimates may be off by 50% or more.

HUPO:
What is HUPO?
The Human Proteome Organization (HUPO) is an international scientific organization representing and promoting proteomics through international cooperation and collaborations by fostering the development of new technologies, techniques and training.

HUPO Mission Statement

To define and promote proteomics through international cooperation and collaborations by fostering the development of new technologies, techniques and training to better understand human disease. View a presentation about HUPOs mission and its initiatives

Objectives

Foster global collaboration in major proteomics projects by gathering leading international laboratories in life sciences, bioinformatics, mass spectrometry, systems biology, pathology, and medicine;

Become the point of contact for proteomics research and commercialization activities worldwide; Support large-scale proteomics projects that are aimed at: A mechanistic understanding of fundamental biological processes (often using model organisms and non human species); Directly studying human disease through proteomics techniques and technologies; Coordinate and enable the fostering of communication among funding agencies and industry partners with the proteomics community and coordinate the activities of groups and organisations interested in HUPOs Scientific Initiatives

Coordinate the development of standard operating procedures related to: Sample preparation, analysis, and repetitions; Data collection, analysis, storage, and sharing; Play a leading role in: Defining the location and functions of proteins in human health and disease by supporting the definition of common and specific standards for peptide and protein characterization from human and model organism specimen selection and phenotypic evaluation to data collection, storage and analysis allowing free and rapid exchange of data; The creation of country-based ethical and legal policy surrounding the handling, banking and use of human tissue specimens for large-scale proteomics projects.

How did HUPO evolve?

HUPO was launched on February 9, 2001. On that date, a global advisory council was officially formed that included leading global experts in the field of proteomics from the academic, government, and commercial sectors. Over the next 12 months, the council, in consultation with industry, identified major proteomics issues and initiatives that needed to be addressed by HUPO. Since its inception, HUPO has received substantial financial assistance from Genome Quebec, Montreal International, McGill University, the National Institutes of Health, and pharmaceutical companies, among others. In addition, it has benefited from considerable in-kind contributions of

time and energy from HUPO Council members, research institutes, and pharmaceutical company partners around the world. Since January 2012, HUPO office is located in Santa Fe, NM. HUPO Initiatives are prominently showcased at each Annual HUPO World Congress, which are held as per a three year rotation in the Americas, Asia/Oceania and Europe. Past congresses have been held in cities such as Versailles, France (2002), Montreal, Canada (2003), Beijing, China (2004), Munich, Germany (2005). Long Beach, USA (2006), Seoul, Korea (2007), Amsterdam, The Netherlands (2008) and Toronto, Canada (2009). The number of participants and exhibitors has significantly increased over the years and the Congresses are a must attend for anyone involved in proteomics.

Arguslab:
ArgusLab is a molecular modeling, graphics, and drug design program for Windows operating systems. Its getting a little dated by now, but remains surprisingly popular. To date, there are > 20,000 downloads. ArgusLab is freely licensed. You dont need to sign anything. You can use as many copies as you need if you are teaching a class where your students might benefit from using ArgusLab. You are not allowed to redistribute ArgusLab from other websites or sources. However, you may link to this website from your own websites if you like. A low-key effort is currently underway to port ArgusLab to the iPad. In addition, Ive done some work with the Qt cross-platform development environment in an effort to support Mac, PC, and Linux.....no promises! :)

Disulphide bonds are formed by the oxidation of two cysteine residues to

form a covalent sulphur-sulphur bond which can be intra- (examples are shown in Jane Richardson's Protein Tourist kinamage) or inter- (exemplified by insulin at this PPS link) molecular bridges. One might imagine that as the enthalpy of a covalent disulphide bond is very high, it contributes a great deal to stability. However, this bond is present in both the folded and the unfolded state, thus its enthalpic contribution to the free energy difference is negligible. All of the stabilizing effect of a disulphide bond is proposed to come from the decrease in conformational entropy of the unfolded state, as described in Conformational Entropy of Unfolding

The Ramachandran Plot

In a polypeptide the main chain N-Calpha and Calpha-C bonds relatively are free to rotate. These rotations are represented by the torsion angles phi and psi, respectively. G N Ramachandran used computer models of small polypeptides to systematically vary phi and psi with the objective of finding stable conformations. For each conformation, the structure was examined for close contacts between atoms. Atoms were treated as hard spheres with dimensions corresponding to their van der Waals radii. Therefore, phi and psi angles which cause spheres to collide correspond to sterically disallowed conformations of the polypeptide backbone.

In the diagram above the white areas correspond to conformations where atoms in the polypeptide come closer than the sum of their van der Waals radi. These regions are sterically disallowed for all amino acids except glycine which is unique in that it lacks a side chain. The red regions correspond to conformations where there are no steric clashes, ie these are the allowed regions namely the alpha-helical and beta-sheet conformations. The yellow areas show the allowed regions if slightly shorter van der Waals radi are used in the calculation, ie the atoms are allowed to come a little closer together. This brings out an additional region which corresponds to the left-handed alpha-helix.

L-amino acids cannot form extended regions of left-handed helix but occassionally individual residues adopt this conformation. These residues are usually glycine but can also be asparagine or aspartate where the side chain forms a hydrogen bond with the main chain and therefore stabilises this otherwise unfavourable conformation. The 3(10) helix occurs close to the upper right of the alpha-helical region and is on the edge of allowed region indicating lower stability. Disallowed regions generally involve steric hindrance between the side chain C-beta methylene group and main chain atoms. Glycine has no side chain and therefore can adopt phi and psi angles in all four quadrants of the Ramachandran plot. Hence it frequently occurs in turn regions of proteins where any other residue would be sterically hindered.
Energy Minimization:

Energy Minimization
Function optimization is a calculation that pervades much of numerical analysis. In the context of macromolecules, the function to be optimized (minimized) is an energy. The energy landscape of a biomolecule possesses an enormous number of minima, or conformational substates. Nonetheless, the goal of energy minimization is simply to find the local energy minimum, i.e., the bottom of the energy well occupied by the initial conformation ( in figure). The energy at this local minimum may be much higher than the energy of the global minimum. Physically, energy minimization corresponds to an instantaneous freezing of the system; a static structure in which no atom feels a net force corresponds to a temperature of 0 K. In the early 1980's, energy minimization was about all one could afford to do and was dubbed `molecular mechanics.'

Energy minimization seeks the energy minimum nearest the starting (marked by star) conformation.