Molecular weight native protein

The shapes of the inhibition curves in Fig. 1A, B, and E are atypical since IgA myeloma proteins are mixtures of monomers and polymers. Using a lower molecular weight dextran N-150N, rather than the higher molecular weight native B512, typical inhibition curves as shown in Fig. 1C were obtained. If the myeloma antidextran was separated into monomer and polymer portions, the usual inhibition curves could be obtained (Cisar et al., 1974). 

FIGURE 8. Determination of the native molecular weight of the purified PSPBP from Acanthocardia tuberculatum foot by native gel electrophoresis of various concentrations of polyacrylamide (6, 8, 10 and 12 %). Molecular weight marker proteins were commercial tetrameric urease (545 kDa), dimeric urease (272 kDa), dimeric BSA (132 kDa), monomeric BSA (66 kDa) and ovalbumin (45 kDa) (Sigma). (A) represents the relative mobilities of proteins plotted as Log (Rf x 100) vs. gel concentration. A plot of the obtained slopes vs. molecular weight was linear and used to determine native PSPBP molecular weight (B). 

A new protein of unknown structure has been purified. Gel filtration chromatography reveals that the native protein has a molecular weight of 240,000. Chromatography in the presence of 6 M guanidine hydrochloride yields only a peak for a protein of M, 60,000. Chromatography in the presence of 6 M guanidine hydrochloride and 10 mM /3-mercaptoethanol yields peaks for proteins of M, 34,000 and 26,000. Explain what can be determined about the structure of this protein from these data. 

The native luciferase having a molecular weight of 106,000 probably consists of two units of the functional 19 kDa protein and two units of the 35 kDa protein. The value of A28o,icm for a solution containing 1 mg/ml of the native luciferase is calculated to be about 0.9 from the inferred amino acid sequence. The function of the 35 kDa protein remains unclear, although it might have a role in the stabilization of the 19 kDa protein. 

A number of different low molecular weight compounds are known to stablize proteins in their native conformation and, therefore, may be effective in correcting of protein folding abnormalities in vivo. Relevant compounds are iV-acetyl-L-lysine, L-camitine, taurine, betaine, ectoine, and hydroxy-ectoine [4]. Some of these chemical chaperones and pharmacological chaperones are already used in clinical trials to combat protein folding diseases, such as cystic fibrosis. 

Figure 14.4 Gel image of proteins extracted from a mixed carbonic anhydrase lysozyme tissue surrogate. Lane M, molecular weight marker lane 1, a 1 2 mol ratio mixture of native, non-formalin-treated carbonic anhydrase and lysozyme lane 2, mixed surrogate with 1 2 mol ratio carbonic anhydrase lysozyme, solubilized and retrieved in 20mM Tris-HCl, pH 4.0, with 2% SDS lane 3, mixed surrogate with 1 2 mol ratio carbonic anhydrase lysozyme, solubilized and retrieved in 20mM Tris-HCl, pH 6.0, with 2% SDS. Protein bands corresponding to lysozyme monomer (a), carbonic anhydrase monomer (b), and the putative lysozyme-carbonic anhydrase heterodimer (c) are indicated. For more detail, see Reference 25.

Figure 15.13 SDS-PAGE of formalin-fixed RNase A before and after protein retrieval. Lane M molecular weight marker lane 1 native RNase A lane 2 formalin-treated RNase A after the removal of excess formaldehyde by rapid dialysis lane 3 formalin-treated RNase A sample from lane 2 after retrieval in 20 mM Tris-HCl, pH 4.0, with 2% SDS lanes 4,6, and 8 formahn-treated RNase A after incubation in 100% ethanol for lh, 24h, or 1 week, respectively lanes 5, 7, and 9 1-h, 24-h, or 1-week formalin-fixed, ethanol-treated RNase A after retrieval in 20 mM Tris-HCl, pH 4.0, with 2% SDS. All RNase A samples were heated at 100°C for 20min, followed by 60°C for 2h. See Fowler et al.12 for details.

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