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Yeast-derived recombinant human insulin-like growth factor I: Production, purification, and structural characterization (1990)

Elliott, Steve ; Fagin, Katherine D. ; Narhi, Linda O. ; [et al.]

Springer

The protein journal 9 (1990), S. 95-104

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ISSN: 1573-4943

Keywords: Insulin-like growth factor ; glycosylation ; disulfide pairing ; circular dichroism ; mutation

Source: Springer Online Journal Archives 1860-2000

Topics: Chemistry and Pharmacology

Notes: Abstract Recombinant human insulin-like growth factor I (IGF-I) is efficiently expressed and secreted fromSaccharomyces cerevisiae using a yeast α-factor leader to direct secretion. However, approximately 10–20% of the IGF-I was in a monomeric form, the remaining materials being disulfide-linked aggregates. When the purified material was subjected to reverse-phase high-performance liquid chromatography (rp-HPLC), it gave two doublet peaks, I and II. Upon reduction, doublet peaks I and II converged to one doublet peak. This suggests that peaks I and II result from different disulfide structures, and the doublet feature of each peak results from other causes. Different disulfide structures between peaks I and II were also suggested from the near UV circular dichroism of these proteins. Only the peak II was biologically active, indicating that peak II has the correct disulfide structure. Concanavalin A affinity chromatography of the purified peak II doublet showed binding of the subpeak with an earlier rp-HPLC retention time, indicating that it was glycosylated. Sequence analysis of tryptic peptides suggested that Thr29 was the site of glycosylation. Site-directed mutagenesis was used to convert Thr29 to Asn29. This substitution reduced, but did not eliminate IGF-I glycosylation, suggesting additional glycosylation sites. The site of carbohydrate addition was consistent with the model that O-glycosylations occur on hydroxyl amino acids near proline residues in β-turns.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1007/BF01024990

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Analysis and measurement of neck loads (1988)

Moroney, Sean P. ; Schultz, Albert B. ; Miller, James A. A.

Hoboken, NJ [u.a.] : Wiley-Blackwell

Journal of Orthopaedic Research 6 (1988), S. 713-720

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ISSN: 0736-0266

Keywords: Neck biomechanics ; Spine biomechanics ; Biomechanical models ; Myoelectric measurements ; Life and Medical Sciences

Source: Wiley InterScience Backfile Collection 1832-2000

Topics: Medicine

Notes: To examine the loads imposed on the structures of the neck by the performance of physical tasks, a biomechanical model of the neck was constructed. The model incorporated 14 bilateral pairs of muscle equivalents crossing the C4 level. A double linear programming optimization scheme that minimized maximum muscle contraction intensity and then vertebral compression force while equilibrating external loads was used to calculate the muscle contraction forces required and the motion segment reactions produced by task performance. To test model validity, 14 healthy adult subjects performed a series of isometric tasks requiring use of their neck muscles. These tasks included exertions in attempted flexion, extension, and left and right lateral bending and twisting. Subjects exerted maximum and submaximum voluntary efforts. During the performance, surface myoelectric activities were recorded at eight locations around the periphery of the neck at the C4 level. Calculated forces and measured myoelectric activities were then linearly correlated. Mean measured voluntary neck strengths in 10 male subjects were as large as 29.7 Nm. Four female subjects developed mean strengths that were approximately 60%-90% of those of the males. In both sexes, neck muscle strengths were approximately one order of magnitude lower than previously measured lumbar trunk strengths. Mean calculated neck muscle contraction forces ranged to 180 N. Mean calculated compression forces on the C4-5 motion segment ranged to 1164 N, lateral shear forces ranged to 125 N, and anteroposterior shear forces ranged to 135 N. Correlation coefficients between the calculated muscle forces and the measured myoelectric activities were as large as 0.85 in some muscles, but generally were smaller than this.

Additional Material: 2 Ill.