Open Access Highly Accessed Open Badges Methodology article

In Silico screening for functional candidates amongst hypothetical proteins

Claus Desler1, Prashanth Suravajhala12, May Sanderhoff1, Merete Rasmussen1 and Lene Juel Rasmussen13*

Author affiliations

1 Department of Science, Systems and Models, Roskilde University, DK-4000 Roskilde, Denmark

2 Centre for Development of Advanced Computing - Bioinformatics Team, Scientific and Engineering Computing Group, Pune University Campus, Pune 411007, India

3 Center for Healthy Aging, Faculty of Health Sciences, Copenhagen University, DK-2200 Copenhagen N, Denmark

For all author emails, please log on.

Citation and License

BMC Bioinformatics 2009, 10:289  doi:10.1186/1471-2105-10-289

Published: 16 September 2009



The definition of a hypothetical protein is a protein that is predicted to be expressed from an open reading frame, but for which there is no experimental evidence of translation. Hypothetical proteins constitute a substantial fraction of proteomes of human as well as of other eukaryotes. With the general belief that the majority of hypothetical proteins are the product of pseudogenes, it is essential to have a tool with the ability of pinpointing the minority of hypothetical proteins with a high probability of being expressed.


Here, we present an in silico selection strategy where eukaryotic hypothetical proteins are sorted according to two criteria that can be reliably identified in silico: the presence of subcellular targeting signals and presence of characterized protein domains. To validate the selection strategy we applied it on a database of human hypothetical proteins dating to 2006 and compared the proteins predicted to be expressed by our selecting strategy, with their status in 2008. For the comparison we focused on mitochondrial proteins, since considerable amounts of research have focused on this field in between 2006 and 2008. Therefore, many proteins, defined as hypothetical in 2006, have later been characterized as mitochondrial.


Among the total amount of human proteins hypothetical in 2006, 21% have later been experimentally characterized and 6% of those have been shown to have a role in a mitochondrial context. In contrast, among the selected hypothetical proteins from the 2006 dataset, predicted by our strategy to have a mitochondrial role, 53-62% have later been experimentally characterized, and 85% of these have actually been assigned a role in mitochondria by 2008.

Therefore our in silico selection strategy can be used to select the most promising candidates for subsequent in vitro and in vivo analyses.