W258–W263 Nucleic Acids Research, 2006, Vol. 34, Web Server issue doi:10.1093/nar/gkl245 i MOD : a powerful and convenient web server for identifying multiple post-translational peptide modifications from tandem mass spectra Sangtae Kim, Seungjin Na 1 , Ji Woong Sim2, Heejin Park3, Jaeho Jeong 4 , Hokeun Kim5, Younghwan Seo6, Jawon Seo4, Kong-Joo Lee4 and Eunok Paek1,* Research Center for Computer Technology, Hanyang University, Seoul, 133-791, Korea, 1Department of Mechanical and Information Engineering, University of Seoul, Seoul 130-743, Korea, 2School of Computer Science and Engineering, Seoul National University, Seoul 151-742, Korea, 3College of Information and Communications, Hanyang University, Seoul 133-791, Korea, 4Center for Cell Signaling Research, Division of Molecular Life Sciences and College of Pharmacy, Ewha Womans University, Seoul 120-750, Korea, 5Functional Proteomics Center, Korea Institute of Science and Technology, Seoul 136-791, Korea and 6Lab Frontier Co. Ltd, Seoul 120-750, Korea Received February 14, 2006; Revised March 14, 2006; Accepted March 29, 2006 ABSTRACT MODi (http://modi.uos.ac.kr/modi/) is a powerful and convenient web service that facilitates the inter- pretation of tandem mass spectra for identifying post-translational modifications (PTMs) in a peptide. It is powerful in that it can interpret a tandem mass spectrum even when hundreds of modification types are considered and the number of potential PTMs in a peptide is large, in contrast to most of the methods currently available for spectra interpretation that limit the number of PTM sites and types being used for PTM analysis. For example, using MODi, one can consider for analysis both the entire PTM list published on the unimod webpage (http://www.unimod.org) and user-defined PTMs simultaneously, and one can also identify multiple PTM sites in a spectrum. MODi is convenient in that it can take various input file for- mats such as .mzXML, .dta, .pkl and .mgf files, and it is equipped with a graphical tool called MassPective developed to display MODi’s output in a user-friendly manner and helps users understand MODi’s output quickly. In addition, one can perform manual de novo sequencing using MassPective. INTRODUCTION Identiﬁcation of post-translational modiﬁcations (PTMs) is important to understand cellular functions of proteins (1). Sensitive methodologies based on conventional biochemical methods are lacking for the identiﬁcation of PTMs in vivo, but recent advances in proteomic technology including mass spectrometry provide an approach to identify PTMs (2–5). Sequencing by tandem mass spectrometry (MS/MS) which has aided protein identiﬁcation offers a tremendous potential for detecting PTMs (1). Three approaches have been used to automatically interpret tandem mass spectra for peptide sequencing (6), namely, data- base searching (7–9), de novo peptide sequencing (10–12) and sequence tag approach (13,14), and there have also been efforts to combine these methods (15,16). Interpretation of experimental spectra is harder if a peptide sequence contains PTMs. One might consider extending one of these approaches in a straightforward manner to sequence a peptide with any number, any kind and any combination of PTMs, i.e. devel- oping a virtual peptide database by incorporating peptides with all possible combinations of PTMs, or extend the set of amino acids by introducing new virtual amino acids such that the mass of each virtual amino acid corresponds to the mass of a post-transitionally modiﬁed amino acid. However, such exten- sions yield an exponential time algorithm or a polynomial time algorithm, the degree of which is very high. Thus these algorithms are not appropriate to interpret tandem mass spec- tra with multiple PTMs in a reasonable amount of time. Recently there have been efforts to formally deﬁne this prob- lem and suggest a way to reduce the time requirement of PTM identiﬁcation (17,18). We have developed a convenient method called MODi for rapidly interpreting tandem mass spectra of peptides with mul- tiple PTMs. This method adopts a hybrid approach that com- bines de novo sequencing with database searching. It performs well even when a large number and types (>100 modiﬁcation *To whom correspondence should be addressed. Tel: +82 2 2210 2680; Fax: +82 2 2248 5110; Email: paek@uos.ac.kr The Author 2006. Published by Oxford University Press. All rights reserved. The online version of this article has been published under an open access model. 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