Recombinant isoforms

Petersen, A., K. Grobe, B. Lindner, M. Schlaak, and W. M. Becker. 1997. Comparison of natural and recombinant isoforms of grass pollen allergens. Electrophoresis 18 (5) 819—825. 

Ex excitation, Em emission, EC extinction coefficient, QY quantum yield, n.d. not determined Recombinant isoforms of Aequorea GFP are described in table 2. 

Table 8. Cloned red fluorescent proteins (RFPs) and recombinant isoforms...

Fig. 4.1.14 Relationship between Ca2+ concentration and the initial light intensity of various recombinant semisynthetic aequorins and w-aequorin J (a semisynthetic natural aequorin made from isoform J). The curve number corresponds to the number of semisynthetic aequorin used in Table 4.1.4. A sample aequorin (3 (Ag) was in 3 ml of calcium-buffer solution containing 1 mM total EGTA, 100 mM KC1,1 mM Mg2+ and 1 mM MOPS (pH 7.0), at 23-24°C. From Shimomura etal., 1993a, with permission from Elsevier.

Masuda, H., et al. (2003). Chromatography of isoforms of recombinant apoaequorin and method for the preparation of aequorin. Protein Expression and Purification 31 181-187. 

Two NKxr splice variants have been identified (Table 3). A NKxr splice variant having a very short C-terminal intracellular tail (7 instead of 96 amino acids), which has been expressed and characterized in recombinant systems (Fig. 1), was found to be expressed at higher level than the long isoform in breast cancer cells. As compared to the long receptor, the short NKxr isoform is less subjected to desensitization and internalization... 

One of the best-studied carrier molecules is produced as a primary excretory constituent of the adult male mouse, known from its consistent high concentration as the major urinary protein (MUP). The basic 3-D structure of the protein was initially obtained from a monoclinic crystal of recombinant protein (MUP-I), constructed by induction in a bacterial expression system and purified to homogeneity (Kuser, 1990). A wild type version of MUP finally yielded to NMR analysis a clone of the r-isoform (162 residues) was labelled and compared with the crystal-structure (Lucke et al., 1990). Two views of the molecule... 

In E. Coli bacterial lysates, the proteome (i.e., the full array of proteins produced) was analyzed by isoelectric focusing and mass spectrometry.97 A comparison of capillary electrophoretic separation and slab gel separation of a recombinant monoclonal antibody demonstrated that the precision, robustness, speed, and ease-of-use of CE were superior.98 Seventy-five proteins from the yeast ribosome were analyzed and identified by capillary electrophoresis coupled with MS/MS tandem mass spectrometry.99 Heavy-chain C-terminal variants of the anti-tumor necrosis factor antibody DE7 have been separated on capillary isoelectric focusing.100 Isoforms differing by about 0.1 pi units represented antibodies with 0,1 or 2 C-terminal lysines. 

Fig. 14.3. Stoichiometry of phosphorylation/inactivation of the bovine kidney and A. suum E1 isoforms. Adult A. suum PDC was depleted of its E1 component and reconstituted with either bovine kidney E1 or recombinant A. suum E1s containing either the al or all isoform (Klingbeil etal., 1997 Huang etal., 1998b). The hybrid complexes were then assayed for PDC activity and the incorporation of 32P, as described fully in Thissen etal. (1986). Upper panel o, bovine kidney E1 , E1 isolated directly from the adult A. suum PDC. Lower panel , A. suum E1al isoform , A. suum E1all isoform.

Burgard, E. C., Tietz, E. I., Neelands, T. R and Macdonald, R. L. (1996) Properties of recombinant y-aminobutyric acidA receptor isoforms containing the as subunit subtype. Mol. Pharmacol. 50, 119-127. 

Although most assays perform well with regard to specificity and reproducibility, the major problem remains their standardization (A9, Dl, K30, L4). There is currently no internationally accepted standard, and the selection of a reference material raised many problems (A8, G5, K30, L4). A number of questions have not been solved Should the standard consist of several apo(a) isoforms Can the reference material be lyophilized Should results be expressed as mass or as moles of apoprotein or lipoprotein How should the protein mass of the primary standard be determined What are optimal storage conditions for the secondary standard Which method can be used as a reference method Can recombinant apo(a) represent an alternative for a primary standard These problems came to light in the course of the international surveys whose results were presented at the Lp(a) Workshop in New Orleans (1992) (L4). 

Table 1. Relative phospholipid specificities for activation of different isoforms of recombinant ATPase II.

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