Vitamin K-dependent γ-glutamylcarboxylase in Atlantic salmon (Salmo salar L.)

Abstract

Due to problems with bone deformities in farmed Atlantic salmon, there is a growing interest in the possible involvement of vitamin K in normal bone development, and sensitive biomarkers for evaluating vitamin K status are therefore needed. The vitamin K-dependent (VKD) enzyme gamma-glutamylcarboxylase (GGCX, EC 6.4.x.x) requires vitamin K as a cofactor for its post-translational modification of glutamic acid (Glu) residues to gamma-carboxyglutamic acid (Gla) residues in VKD proteins, and is required for their function in haemostasis and bone metabolism. The present study was designed to evaluate the enzyme assay for GGCX activity in isolated liver microsomes and its distribution in the tissues of Atlantic salmon. The effect of KH(2) and menadione on the GGCX activity in salmon liver was also compared. Results from the present study show a widespread tissue distribution and expression of GGCX in Atlantic salmon. The GGCX activity and ggcx expression in all bony tissues examined imply the presence of vitamin K, and suggest the involvement of vitamin K in bone metabolism of Atlantic salmon. We propose the GGCX assay as a sensitive marker for vitamin K status, and confirm that menadione does not work as a cofactor for GGCX in Atlantic salmon liver.

Vitamin K in fish nutrition

“…Menadione must be alkylated enzymatically to MK‐4 in animal tissues to become biologically active (Udagawa 2000), and is thought to have lower bioavailability than the naturally occurring K vitamers, as shown in mummichog (Udagawa 2001; Udagawa & Murai 2001). Results from Krossøy et al. (2010) confirmed that menadione did not act directly as a co‐factor for GGCX in Atlantic salmon, despite its structural similarities with the inherent vitamin K forms (Lambert & De Leenher 1992).…”

“…As the major function of vitamin K is to act as co‐factor for GGCX, the activity of this enzyme may provide a biomarker for deficiency. Results from recent studies in juvenile Atlantic salmon confirmed that GGCX activity is a sensitive marker for evaluating vitamin K status and intake (Krossøy et al. 2009a, 2010).…”

“…2009a,b). Furthermore, gene expression of ggcx was found in vertebrae, scales, operculum and fin of adult Atlantic salmon, indicating GGCX activity in bony tissues of Atlantic salmon (Krossøy et al. 2010).…”

Higher dietary micronutrients are required to maintain optimal performance of Atlantic salmon (Salmo salar) fed a high plant material diet during the full production cycle

“…Menadione is very unstable during feed processing (Krossøy et al, 2009), and the analysed dietary levels of K3 were only 1/10 of the supplemented levels. Menadione is metabolised to menakinones (MK4-13, vitamin K2) in the body (Krossøy et al, 2011) and therefore, MK4 increased in response to increase in dietary vitamin K3, while whole body vitamin K1 was similar in fish fed the different treatments. Krossøy et al (2009) fed Atlantic salmon eight diets supplemented with 0 to 50 mg kg -1 vitamin K3 and found no effects on fish performance, or blood coagulation time and vertebrae stiffness.…”

Enhanced micronutrient supplementation in low marine diets reduced vertebral malformation in diploid and triploid Atlantic salmon (Salmo salar) parr, and increased vertebral expression of bone biomarker genes in diploids

“…Many of the micronutrients act as important cofactors in enzymatic pathways and other metabolic processes in vertebral formation. Several studies in diploid salmon have demonstrated a key role of nutrients, particularly P and Ca (Åsgård and Shearer, 1997;Baeverfjord et al, 1998), vitamin A (Green et al, 2016) and vitamin K (Krossøy et al, 2010), in skeletal development and either their deficiency or toxicity leads to pathogenesis of skeletal deformities. Vitamin A regulates skeletogenesis and cartilage development by controlling chondrocyte function, maturation and proliferation of cells (Koyama et al, 1999), while vitamin K…”