Catalytic nucleotide-hydrolyzing antibodies in milk and serum of clinically healthy human mothers.
BACKGROUND
In humans, pregnancy and lactation are associated with the production of catalytically active antibodies (abzymes) in serum and breast milk. However, the substrate specificities of the abzymes in these biological fluids, particularly breast milk, have not been studied


METHODS
IgG fractions were isolated from human milk by subsequent steps of chromatographic purification on Protein-A Sepharose, DEAE-cellulose, and anti-IgG Sepharose. The nucleotide-hydrolyzing activity of electrophoretically homogeneous IgG antibodies was measured using 32P-labeled nucleotides and TLC.


RESULTS
We demonstrated by different methods that IgG antibodies from the serum and milk of clinically healthy human mothers are able to hydrolyze ribo- and deoxyribonucleoside-5'-mono, di- and triphosphates; this nucleotide-hydrolyzing activity was also present in Fab fragments of the IgG molecule. Affinity modification of the milk IgG oligomeric form by chemically reactive derivatives of ATP led to preferential modification of the L-chain. However, after separation of the subunits by SDS electrophoresis, an in-gel assay showed ATP-hydrolyzing activity in various oligomeric forms of IgG subunits (H2L2, H2L and HL), while the separated heavy (H) and light (L) chains were not catalytically active. The Km and Vmax values characterizing the interaction of IgG with nucleotides were estimated.


CONCLUSIONS
Our findings speak in favor of the generation of a variety of polyclonal nucleotide-hydrolyzing antibodies by the immune system of clinically healthy mothers.
Semenov
Dmitry V
DV
Novosibirsk Institute of Bioorganic Chemistry, Siberian Division of the Russian Academy of Sciences, Novosibirsk 630090, Lavrientieva Avenue 8, Russia.
Kanyshkova
Tat'yana G
TG
Karotaeva
Nataliya A
NA
Krasnorutskii
Mikhail A
MA
Kuznetsova
Irina A
IA
Buneva
Valentina N
VN
Nevinsky
Gregory A
GA
D000249:Adenosine Monophosphate / Q000031:analogs & derivatives / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D000328:Adult; D017151:Antibodies, Catalytic / Q000097:blood / Q000302:isolation & purification / Q000378:metabolism*; D005260:Female; D006801:Humans; D006868:Hydrolysis; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D007074:Immunoglobulin G / Q000302:isolation & purification / Q000378:metabolism*; D007143:Immunoglobulin Heavy Chains / Q000378:metabolism; D007147:Immunoglobulin Light Chains / Q000378:metabolism; D007700:Kinetics; D007774:Lactation; D008895:Milk, Human / Q000737:chemistry / Q000276:immunology*; D009711:Nucleotides / Q000737:chemistry / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D011247:Pregnancy; D044967:Serum / Q000737:chemistry*
D017151:Antibodies, Catalytic; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D009711:Nucleotides; D010761:Phosphorus Radioisotopes; D000249:Adenosine Monophosphate; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases