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Gene synthesis, high‐level expression and assignment of backbone 15 N and 13 C resonances of soybean leghemoglobin
Author(s) -
Prytulla Stefan,
Jane Dyson H.,
Wright Peter E.
Publication year - 1996
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(96)01278-1
Subject(s) - leghemoglobin , gene expression , chemistry , physics , gene , biology , genetics , nitrogen fixation , bacteria , root nodule
A synthetic gene for apoleghemoglobin‐a from soybean, optimized for expression in Escherichia coli has been designed and synthesized by a recursive polymerase chain reaction technique. The protein has been expressed with high efficiency and a purification protocol has been developed. The holoprotein is readily reconstituted by the addition of heme. 15 N‐ and 15 N, 13 C‐labeled samples were produced and backbone 15 N and 13 C assignments were determined by 2D and 3D NMR spectroscopy. Comparison of the chemical shifts of 13 C α and 13 CO with random coil shifts revealed a pattern of secondary structure which correlates well with the one previously derived from homonuclear NMR data and low‐resolution X‐ray crystallography.