This article describes the development of a new method for multiple sequence alignment based on fold-level protein structure alignments, which provides an improvement in accuracy compared with the most commonly used sequence-only-based techniques. This method integrates the widely used, progressive multiple sequence alignment approach ClustalW with the Topology of Protein Structure (TOPS) topology-based alignment algorithm. The TOPS approach produces a structural alignment for the input protein set by using a topology-based pattern discovery program, providing a set of matched sequence regions that can be used to guide a sequence alignment using ClustalW. The resulting alignments are more reliable than a sequence-only alignment, as determined by 20-fold cross-validation with a set of 106 protein examples from the CATH database, distributed in seven superfold families. The method is particularly effective for sets of proteins that have similar structures at the fold level but low sequence identity. The aim of this research is to contribute towards bridging the gap between protein sequence and structure analysis, in the hope that this can be used to assist the understanding of the relationship between sequence, structure and function. The tool is available at http://balabio.dcs.gla.ac.uk/msat/.