title: Structure of iron saturated C-lobe of bovine lactoferrin at pH 6.8 indicates a weakening of iron coordination.
abstract: The bilobal lactoferrin is an approximately 76 kDa glycoprotein. It sequesters two Fe(3+) ions together with two CO(3)(2-) ions. The C-terminal half (residues, Tyr342-Arg689, C-lobe) of bovine lactoferrin (BLF) (residues Ala1-Arg689) was prepared by limited proteolysis using trypsin. Both C-lobe and intact BLF were saturated to 100%. Both of them retained up to nearly 85% of iron at pH 6.5. At pH 5.0, C-lobe retained 75% of iron whereas intact protein could retain only slightly more than 60%. At pH 4.0 both contained 25% iron and at pH 2.0 they were left with iron concentration of only 10%. The structure of iron saturated C-lobe was determined at 2.79 Å resolution and refined to R(cryst) and R(free) factors of 0.205 and 0.273, respectively. The structure contains two crystallographically independent molecules, A and B. They were found to have identical structures with an r.m.s. shift of 0.5 Å for their C(α) atoms. A high solvent content of 66% was observed in the crystals. The average value of an overall B-factor was 68.0 Å(2). The distance of 2.9 Å observed for the coordination bond between Fe(3+) ion and N(e2) of His595 appeared to be considerably longer than the normally observed values of 1.9-2.2 Å. This indicated that the coordination bond involving His595 may be absent. Other coordination distances were observed in the range of 2.1-2.3 Å. Based on the present structure of iron saturated C-lobe, it may be stated that His595 is the first residue to dissociate from ferric ion when the pH is lowered.
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Nilisha
authors: N
authors: Rastogi
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Avinash
authors: A
authors: Singh
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Prashant K
authors: PK
authors: Singh
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Tapesh K
authors: TK
authors: Tyagi
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Sada
authors: S
authors: Pandey
authors: Functional Food Ingredients Department, Morinaga Milk Industry Co. Ltd, Kanagawa, Japan.
authors: Kouichirou
authors: K
authors: Shin
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Punit
authors: P
authors: Kaur
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Sujata
authors: S
authors: Sharma
authors: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors: Tej P
authors: TP
authors: Singh
mesh_terms: D000818:Animals; D002417:Cattle; D006863:Hydrogen-Ion Concentration; D007501:Iron / Q000737:chemistry* / Q000378:metabolism*; D007781:Lactoferrin / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D014357:Trypsin / Q000378:metabolism
chemical_list: D010446:Peptide Fragments; D007501:Iron; D007781:Lactoferrin; D014357:Trypsin
keywords: C-lobe; lactoferrin; structure; tryptic digestion; weak iron coordination