Crystallization and preliminary X-ray analysis of W120K mutant of streptavidin.
Bacterial streptavidin and chicken avidin are homotetrameric proteins that share an exceptionally high affinity towards the vitamin biotin. The biotin-binding sites in both proteins contain a crucial tryptophan residue contributed from an adjacent subunit. This particular tryptophan (W110 in avidin and W120 in streptavidin) plays an important role in both biotin binding and in the quaternary stabilities of the proteins. An intriguing naturally occurring alteration of tryptophan to lysine was previously described in the C-terminal domain of sea-urchin fibropellins, which share a relatively high sequence similarity with avidin and streptavidin. Avidin (Avm-W110K) and streptavidin (Savm-W120K) mutations show substantially reduced affinities towards biotin as well as the dissociation of their tetrameric structure into stable avidin and streptavidin dimers. Savm-W120K was crystallized at 293 K using the hanging-drop vapour-diffusion method. The crystals diffract to 1.7 A resolution using synchrotron radiation and belong to the monoclinic space group P2(1), with unit-cell parameters a = 50.43, b = 100.41, c = 52.51 A, beta = 112.12 degrees. The asymmetric unit contains four molecules of Savm-W120K, with a corresponding V(M) of 2.3 A(3) Da(-1) and a solvent content of 46%.
Pazy
Y
Y
Department of Biological Chemistry, The Institute of Life Sciences, The Wolfson Centre for Applied Structural Biology, The Hebrew University of Jerusalem, Givat Ram, Jerusaleum 91904, Israel.
Laitinen
O H
OH
Ravoy
B
B
Kulomaa
M S
MS
Wilchek
M
M
Bayer
E A
EA
Livnah
O
O
D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry*; D016367:Baculoviridae / Q000235:genetics; D003460:Crystallization; D018360:Crystallography, X-Ray; D008239:Lysine / Q000235:genetics; D009154:Mutation; D011487:Protein Conformation; D019809:Streptavidin / Q000737:chemistry* / Q000235:genetics; D014364:Tryptophan / Q000235:genetics
D001426:Bacterial Proteins; D014364:Tryptophan; D019809:Streptavidin; D008239:Lysine