A possible approach to generate enzymes with an engineered temperature optimum is to create chimeras of homologous enzymes with different temperature optima. We tested this approach using two family-10 xylanases from Thermotoga maritima: the thermophilic xylanase A catalytic domain (TmxAcat, T-opt = 68 degrees C), and the hyperthermophilic xylanase B (TmxB, T-opt = 102 degrees C). Twenty-one different chimeric constructs were created by mimicking family shuffling in a rational manner. The measured temperature optima of the 16 enzymatically active chimeras do not monotonically increase with the percentage of residues coming from TmxB. Only four chimeras had a higher temperature optimum than TmxAcat, the most stable variant (T-opt = 80 degrees C) being the one in which both terminal segments came from TmxB. Further analysis suggests that the interaction between the N- and C-terminal segments has a disproportionately high Contribution to the overall thermostability. The results may be generalizable to other enzymes where the N- and C-termini are in contact. (c) 2008 Elsevier Inc. All rights reserved.