A 44-kDa of protein identical to the N-terminal amino acid sequence of MCT1 in human circulation.
A family of specific carrier protein designated as monocarboxylate transporter (MCT) has been known to transport the lactate and other moncarboxylates in mammalian cells. We hypothesized the presence of serum protein in human circulation that may works as a lactate carrier and that biochemical structure would possesses common structure with MCT on the plasma membrane. Immunoblot analysis with an anti-MCT1 polyclonal antibody suggested the presence of a 44-kDa protein in human circulation and N-terminal amino acid sequencing exhibited a stretch of 14 amino acids which is completely identical to MCT1. The unbound fractions from the GST-MCTI fusion protein-immobilized glutathione sepharose 4B column demonstrated that lactic acid concentration began to increase with one fraction delay compared to Sepharose 4B and GST-immobilized column. When lactic acid was washed away with PBS, lactic acid concentrations in the effuluent constantly decreased in both Sepharose 4B and GST-immobilized column. However, GST-MCT1-immobilized column showed specific convex curve from fraction approximately 3 mM of lactate and demonstrated wash out delay compared to Sepharose 4B and GST-immobilized column. These observations demonstrated biochemical and immunological similarities between a 44-kDa protein purified from human serum and MCT1 present on the plasma membrane. The studies on MCT1-fusion protein suggested possible functional properties of a 44-kDa protein as a lactate buffer by holding and unhand a lactate according to the lactate concentration in human blood. The experiments described herein have suggested the existence of lactate carrier in human circulation which is free from plasma membrane.
Iizuka
Kenji
K
Department of Laboratory Medicine, Hokkaido University Graduate School of Medicine, Sapporo, Japan. kiizuka@med.hokudai.ac.jp
Morita
Noriteru
N
Nagai
Tatsuya
T
Hanada
Akiko
A
Okita
Koichi
K
Yonezawa
Kazuya
K
Murakami
Takeshi
T
Kitabatake
Akira
A
Kawaguchi
Hideaki
H
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D018076:DNA, Complementary / Q000378:metabolism; D004912:Erythrocytes / Q000378:metabolism; D015723:Gene Library; D005982:Glutathione Transferase / Q000378:metabolism; D006352:Heart Ventricles / Q000473:pathology; D006801:Humans; D015151:Immunoblotting; D019344:Lactic Acid / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins; D019344:Lactic Acid; D005982:Glutathione Transferase