A complex family of highly heterogeneous and internally repetitive hyperactive antifreeze proteins from the beetle Tenebrio molitor.
We have previously identified a Thr- and Cys-rich thermal hysteresis (antifreeze) protein (THP) in the beetle Tenebrio molitor that has 10-100 times the freezing point depression activity of fish antifreeze proteins. Because this 8.4 kDa protein is significantly different in its properties from THP preparations previously reported from this insect, a thorough search was undertaken for other antifreeze types. Many active proteins were observed, but all appeared to be isoforms of the THP that differed in their number of 12-amino acid repeats (consensus sequence CTxSxxCxxAxT), amino acid substitutions, and N-linked glycosylation. Mass spectral analysis has matched most of these isoforms with cDNA sequences of 17 different clones from a larval fat body library that encode eight different mature THPs containing 84, 96, or 120 amino acids. Genomic Southern blots suggest there may be 30-50 tightly linked copies of the gene, which is a signature consistently seen with unrelated fish antifreeze protein genes, and one that has been associated with the need to rapidly increase gene product in response to climate change. A three-dimensional model is proposed for the fully disulfide-bonded structure of T. molitor THP, which can accommodate addition or deletion of 12-amino acid repeats. The structure is a beta-helix that places most of the Thr in a regular array on one side of the protein to form a putative ice-binding surface.
Liou
Y C
YC
Department of Biochemistry, Queen's University, Kingston, Ontario, Canada.
Thibault
P
P
Walker
V K
VK
Davies
P L
PL
Graham
L A
LA
D000595:Amino Acid Sequence; D000818:Animals; D021301:Antifreeze Proteins; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000302:isolation & purification; D005615:Freezing; D017344:Genes, Insect; D006023:Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D006458:Hemolymph / Q000378:metabolism; D006358:Hot Temperature; D019476:Insect Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D005810:Multigene Family; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D020449:Repetitive Sequences, Amino Acid; D013712:Tenebrio / Q000737:chemistry* / Q000235:genetics*
D021301:Antifreeze Proteins; D018076:DNA, Complementary; D006023:Glycoproteins; D019476:Insect Proteins; D020033:Protein Isoforms