Abstract
The `phase problem' in crystallography results from the inability to directly measure the phases of individual diffracted X-ray waves. While intensities are directly measured during data collection, phases must be obtained by other means. Several phasing methods are available (MIR, SAR, MAD, SAD and MR) and they all rely on the premise that phase information can be obtained if the positions of marker atoms in the unknown crystal structure are known. This paper is dedicated to the most popular phasing method, molecular replacement (MR), and represents a personal overview of the development, use and requirements of the methodology. The first description of noncrystallographic symmetry as a tool for structure determination was explained by Rossmann and Blow [Rossmann & Blow (1962), Acta Cryst. 15, 24-31]. The term `molecular replacement' was introduced as the name of a book in which the early papers were collected and briefly reviewed [Rossmann (1972), The Molecular Replacement Method. New York: Gordon & Breach]. Several programs have evolved from the original concept to allow faster and more sophisticated searches, including six-dimensional searches and brute-force approaches. While careful selection of the resolution range for the search and the quality of the data will greatly influence the outcome, the correct choice of the search model is probably still the main criterion to guarantee success in solving a structure using MR. Two of the main parameters used to define the `best' search model are sequence identity (25% or more) and structural similarity. Another parameter that may often be undervalued is the quality of the probe: there is clearly a relationship between the quality and the correctness of the chosen probe and its usefulness as a search model. Efforts should be made by all structural biologists to ensure that their deposited structures, which are potential search probes for future systems, are of the best possible quality.
Highlights
The original concept of molecular replacement (MR) was introduced in the early 1960s; the first paper mentioning a way to solve a crystal structure using a search model was likely to have been a paper by Huber (1965)
There have been two previous CCP4 Study Weekends devoted to MR
Molecular replacement has come a long way from the initial programs in accuracy, speed, ease of use and level of automation
Summary
The original concept of molecular replacement (MR) was introduced in the early 1960s; the first paper mentioning a way to solve a crystal structure using a search model was likely to have been a paper by Huber (1965). The growth is exponential, and it is even more compelling if we look at the number of structures solved using MR compared with the entire body of X-ray-derived structures (Fig. 1b): overall, almost 60% of the structures have been solved using MR, and in the past two years they account for over 70% of all depositions This impressive growth has been made possible by a dramatic evolution of both computer hardware and software, which has resulted in a faster, more flexible and in many cases fully automated methodology, and a dramatic evolution and improvement in the type, quantity and quality of the starting models. This paper aims to be a review of the development of the method, with examples taken from the literature and the author’s own experience
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