M Tomita, N Ohkura, M Ito, T Kato, PM Royce, T Kitajima
BIOCHEMICAL JOURNAL 312 847-853 1995年12月
We have investigated the expression of human procollagen III by insect cells infected with a recombinant baculovirus carrying cDNA for the pro-alpha 1(III) chain of type-III collagen. A high level of expression was obtained, and a small proportion of the heterologously expressed pro-alpha 1(III) chains formed normally disulphide-bonded procollagen III, which was secreted into the culture medium. This species displayed a melting temperature (T-m) of approx. 38 degrees C as assessed by its resistance to digestion by a mixture of trypsin and chymotrypsin, slightly lower than that of 39.5 degrees C for procollagen III synthesized by cultured human dermal fibroblasts, and reflected a slight degree of under-hydroxylation of prolyl residues. This is possibly a consequence of the lower incubation temperature of insect cells, or of an insufficiency of prolyl hydroxylase activity within them. A significant proportion of the expressed chains formed trimeric molecules of similar thermal stability containing an apparently full-length triple-helical region, but were not disulphide-bonded and not secreted. In addition to providing a source of recombinant human procollagen III, the system promises to be useful in the study of procollagen chain association and subsequent folding.
T OKA, Y ENDO, M ITO, K MIYAMOTO, T SASAKAWA, SUZUKI, I, Y NATORI
BIOCHIMICA ET BIOPHYSICA ACTA 1130(2) 224-226 1992年3月
The cDNA clone encoding HMG 2a of chick liver was isolated from a lambda-gt11 expression library using polyclonal antibodies. DNA sequence analysis revealed an open reading frame of 201 amino acids. Comparison of the nucleotide sequences of cDNA coding for chick liver HMG 2a with pig thymus HMG 2 and human monocytic leukemia cell HMG 2 showed 70% homology. 2.0 kb and 1.2 kb mRNAs were found in newly hatched chick liver and decreased during postnatal development of chicks.